The effect and mechanisms of risperidone and voluntary exercise intervention on hepatic lipid metabolism in juvenile female rats

The effect and mechanisms of risperidone and voluntary exercise intervention on hepatic lipid metabolism in juvenile female rats | Research Square window.SnipcartSettings = { analytics: { enabled: false } }; (function() { var accessVector = localStorage.getItem('access_vector') || ''; window.dataLayer = window.dataLayer || []; if (accessVector) { window.dataLayer.push({ user: { profile: { profileInfo: { snid: accessVector } } } }); } })(); (function(w,d,s,l,i){w[l]=w[l]||[];w[l].push({'gtm.start':new Date().getTime(),event:'gtm.js'});var f=d.getElementsByTagName(s)[0],j=d.createElement(s),dl=l!='dataLayer'?'&l='+l:'';j.async=true;j.src='https://www.googletagmanager.com/gtm.js?id='+i+dl;f.parentNode.insertBefore(j,f);})(window,document,'script','dataLayer','GTM-K279D39R'); Browse Preprints In Review Journals COVID-19 Preprints AJE Video Bytes Research Tools Research Promotion AJE Professional Editing AJE Rubriq About Preprint Platform In Review Editorial Policies Our Team Advisory Board Help Center Sign In Submit a Preprint Cite Share Download PDF Research Article The effect and mechanisms of risperidone and voluntary exercise intervention on hepatic lipid metabolism in juvenile female rats Weijie Yi, Jiamei Lian, Chao Deng This is a preprint; it has not been peer reviewed by a journal. https://doi.org/ 10.21203/rs.3.rs-7309963/v1 This work is licensed under a CC BY 4.0 License Status: Posted Version 1 posted You are reading this latest preprint version Abstract Objectives Risperidone is a commonly used antipsychotic drug in juveniles, but with serious metabolic side-effects. Previous studies found that exercise reduced plasma triglyceride level and adipose accumulation caused by risperidone. This study elucidated the underlying mechanisms. Methods Female juvenile rats were randomly allocated into Vehicle + Sedentary, Risperidone (0.9mg/kg; twice per day) + Sedentary, Vehicle + Exercise (3-hour voluntary access to a running wheel/day), and Risperidone + Exercise groups (n = 8/group). After 4-week treatment, the liver was harvested for subsequent examination. Results (1) Lipogenesis: Protein levels of FAS and USF1 were raised in the risperidone-treated sedentary group, which was decreased by exercise. The pAMPK/AMPK ratio was upregulated by exercise. (2) Lipid uptake/storage: Risperidone-induced upregulations of PPARγ and CD36 were downregulated by exercise. FSP27 expression was decreased by exercise. (3) Lipolysis/β-oxidation: Hepatic protein levels of ATGL and HSL in the Risperidone + Exercise group were larger than Risperidone + Sedentary group. Reduced PGC1α expression was found in the risperidone-only group, which was reversed by exercise. Conclusion Risperidone enhanced fatty acid synthesis via the hepatic USF1/FAS signaling pathway and to augment fatty acid uptake through the PPARγ/CD36 pathway, while simultaneously diminishing β-oxidation by down-regulating hepatic PGC1α expression. Conversely, voluntary exercise intervention counteracted these effects, thereby ameliorating the lipid imbalances induced by risperidone. Antipsychotic drug Exercise Metabolic side effect Lipid metabolism Juvenile Figures Figure 1 Figure 2 Figure 3 Introduction Risperidone (neuroscience-based nomenclature (NbN): [dopamine D2, serotonin 5-HT2, and norepinephrine alpha-2 receptor antagonist]), one of the most commonly prescribed second-generation antipsychotics (SGAs), accounts for approximately 70% of antipsychotic prescriptions in juveniles under 14 years of age (Klau et al., 2024 ). However, it is associated with significant metabolic side effects, including weight gain, insulin resistance, and dyslipidemia, which can lead to metabolic syndrome (Pillinger et al., 2020 ). Vulnerable populations such as children, adolescents, and females are particularly susceptible to these adverse effects (Castellani et al., 2019 ; Morrato et al., 2010 ). A prospective study reported that more than half (53.8%) of pediatric patients receiving risperidone experienced at least one metabolic abnormality, with hyperlipidemia being the most common (34.6%)(Alsabhan et al., 2024 ). Additionally, female sex, considered both a risk factor and a predictive marker for SGA-induced weight gain, likely reflects increased vulnerability due to sex-specific physiological characteristics, such as a higher proportion of adipose tissue and the modulatory influence of gonadal hormones(Fitzgerald et al., 2003 ; Gebhardt et al., 2009 ; Kelly et al., 1999 ). Voluntary exercise has been shown to improve lipid metabolism (Wooten et al., 2022 ). In our previous work, we found that voluntary exercise significantly attenuated risperidone-induced increases in plasma triglyceride levels and adipose tissue accumulation in juvenile rats (Yi et al., 2021 ). However, the underlying mechanisms responsible for these protective effects remain incompletely understood. The liver is essential in maintaining whole-body lipid homeostasis by regulating the synthesis, storage, modification, and transport of lipids. Hepatic de novo lipogenesis contributes to the storage and secretion of lipids from hepatocytes (Jensen-Urstad and Semenkovich, 2012 ). Insulin activates lipogenic transcription factors [sterol regulatory element binding transcription factor 1c (SREBP1c), liver X receptor (LXR), and upstream transcription factor 1 (USF1)] upregulate the expression of lipogenic enzymes [e.g. Fatty acid synthase (FAS), Acetyl-CoA carboxylase1 (ACC1) and Stearoyl-CoA desaturase (SCD1)], resulting in fatty acid synthesis (Liu et al., 2020b ). Synthesized fatty acids are stored in the liver on the form of triglycerides and exported into the bloodstream in very-low-density lipoprotein (VLDL) particles. Additionally, fatty acids could be released from triglycerides through the catalysis of adipose triglyceride lipase (ATGL) and hormone-sensitive lipase (HSL), and then plasma fatty acids are taken up into the liver by fatty acid transport protein 2 (FATP2), caveolin-1 (CAV1) and cluster of differentiation 36 (CD36). Moreover, peroxisome proliferator-activated receptor α (PPARα) and γ (PPARγ) also modulate fatty acid uptake, trafficking, catabolism, utilization, triglyceride synthesis, and lipid droplet formation (Dong et al., 2024 ; Su et al., 2020 ). Further, the majority of the fatty acids in hepatocytes is translocated into the mitochondria and undergo β-oxidation (Alves-Bezerra and Cohen, 2017 ). Carnitine palmitoyltransferase 1A (CPT1A), a downstream target of PPARα and a rate-limiting enzyme for fatty acid β-oxidation, facilitating fatty acids entering the mitochondrial matrix (Houten et al., 2016 ). Peroxisome proliferator-activated receptor-γ coactivator 1-α (PGC-1α) collaborates with PPARα to regulate the expression of fatty acid oxidation enzymes in mitochondria (Vega et al., 2000 ). Imbalances between lipid synthesis and degradation leads to lipid metabolism disorders. Treatment with SGAs has been reported to disrupt hepatic lipogenesis, lipolysis, fatty acids uptake, and β-oxidation (Oh et al., 2011 ; Su et al., 2023 ). Meanwhile, exercise improves lipid homeostasis by reducing synthesis and transport of fatty acids triglyceride in both adipose tissue and liver (Kurosaka et al., 2021 ; May et al., 2017 ). To date, no study has investigated the mechanisms through which exercise ameliorates lipid metabolism disorders induced by risperidone. Therefore, this study explored the possible mechanisms driving the effects of voluntary exercise in alleviating risperidone-induced lipid metabolic disorders in a juvenile female rat model. Materials and Methods Animal housing and treatment Animal housing and treatment protocols were conducted as previously detailed (Yi et al., 2021 ). Briefly, juvenile female Sprague-Dawley rats (postnatal day 22/23) were obtained from the Animal Resource Centre (Perth, Western Australia). At postnatal day 26/27, they were housed individually in Techniplast GR1800 ventilated cage (Lane Cove West, NSW, Australia), and randomly allocated into (1) Vehicle + Sedentary (VS), (2) Risperidone + Sedentary (RS), (3) Vehicle + Exercise (VE), and (4) Risperidone + Exercise (RS) groups (n = 8/group). Risperidone (0.9 mg/kg; Janssen, Macquarie Park, NSW, Australia)was administered at a total dose of 1.8 mg/kg/day (0.9 mg/kg per dose, twice daily at 07:00 and 19:00) in 0.3 g cookie dough pellets from postnatal days 29/30 for a duration of 4 weeks, while the control rats were given plain cookie dough pellets (15% gelatine, 9% milk powder, 38% corn flour and 38% sugar) at the same times. The dosage was translated from the clinical dose based on body surface area in accordance with FDA guidelines (FDA, 2005 ; Reagan-Shaw et al., 2008 ), and has been shown to be physiologically and behaviorally effective in juvenile rats (De Santis et al., 2016 ; De Santis et al., 2018 ; Lian et al., 2015 ; Sylvester et al., 2020 ). On postnatal day 57/58, following overnight fasting, the final dose of risperidone was administered orally using a 1 mL syringe, with the drug dissolved in approximately 0.2 mL of water to avoid the potential impact of cookie dough on plasma glucose and lipid levels. Rats were allowed to voluntarily access running wheels equipped with revolution counters for 3 hours daily in a 4-week period (from postnatal days 29/30 to 56/57), with traveling distance recorded (Scurry Rat Running Wheel/Chamber, Lafayette Instrument, IN, USA). All rats were euthanized by decapitation following isoflurane anesthesia on postnatal day 57/58. Tissue samples (liver, inguinal, perirenal, periovary, and mesentery adipose tissue) were harvested and weighed immediately, and then frozen in liquid nitrogen and kept at -80°C. Blood was collected from the left ventricle into EDTA tube, and the plasma was separated by centrifuge (4℃, 3000 rpm, 10 min) then stored at -80°C until further use. As previously reported, Risperidone treatment significantly reduced physical activity over the 28-day intervention period (Average distance travelled: RE group, 1656.13 ± 359.03 m/day vs VE group, 2828.00 ± 416.24 m/day, p < 0.05) (Yi et al., 2021 ). Voluntary exercise reduced risperidone-induced increases in adipose tissue [periovary index (VS: 0.89 ± 0.06, RS: 1.27 ± 0.12, VE: 0.75 ± 0.06, RE: 0.85 ± 0.13), perirenal index (VS: 0.84 ± 0.09, RS: 1.06 ± 0.10, VE: 0.63 ± 0.04, RE: 0.78 ± 0.10) and inguinal (VS: 1.15 ± 0.09, RS: 1.56 ± 0.08, VE: 1.01 ± 0.05, RE: 1.25 ± 0.11)], fasting plasma insulin (VS: 85.27 ± 6.35, RS: 201.16 ± 48.08, VE: 129.27 ± 19.37, RE: 113.67 ± 16.76 pmol/L), and triglycerides(VS: 0.67 ± 0.09, RS: 1.22 ± 0.18, VE: 0.56 ± 0.06, RE: 0.71 ± 0.08 mM) (Yi et al., 2021 ). The Animal Ethics Committee, University of Wollongong, Australia, approved all experimental procedures (AE18/19). Western blots Procedures of the liver lysate preparations and Western blot were conducted as reported previously (Yi et al., 2021 ). In brief, aliquots containing 15 µg protein were added to electrophoresis on a precast polyacrylamide (4–20%) gel (Bio-Rad Laboratories, Gladesville, NSW, Australia). transferred the separated protein to a polyvinylidene difluoride (PVDF) membrane (Bio-Rad Laboratories, Gladesville, NSW, Australia). Following transferred the separated protein to a polyvinylidene difluoride membrane, it was blocked with 5% skim milk plus 0.1% Tween-20 in Tris-buffered saline for a subsequent overnight incubation at 4°C with following primary antibodies: anti-SREBP1 (1:500, #ab28481, Abcam, Cambridge, UK), anti-SCD1 (1:1000, #ab19862, Abcam, Cambridge, UK), anti-PPARγ (1:1000, ab209350, Abcam, Cambridge, UK), anti-USF1 (1:1000, #ab180717, Abcam, Cambridge, UK), anti-ATGL (1:1000, #ab109251, Abcam, Cambridge, UK), anti-FSP27 (1:1000, #ab213693, Abcam, Cambridge, UK), anti-PGC1α (1:1000, #ab191838, Abcam, Cambridge, UK), anti-LXRα(1:1000, #ab106464, Abcam, Cambridge, UK), CAV-1(1:1000, #ab2910, Abcam, Cambridge, UK) HSL(1:1000, #ab45422, Abcam, Cambridge, UK), FABP1 (1:1000, #ab222517, Abcam, Cambridge, UK), anti-FAS (1:1000, #3180S, Cell Signaling, Danvers, MA, USA), anti-CD36 (1:1000, #74002, Cell Signaling, Danvers, MA, USA), anti-SCAP (1:1000, #13102S, Cell Signaling, Danvers, MA, USA), anti-pAMPKα(1:2000, #2535S, Cell Signaling, Danvers, MA, USA), anti-AMPKα(1:1000, #2532, Cell Signaling, Danvers, MA, USA), anti-ACC (1:500, #3662S, Cell Signaling, Danvers, MA, USA) anti-INSIG2 (1:1000, #PA5109863, Invitrogen, Camarillo, USA), anti-FATP2 (1:1000, #MA5-50447, Invitrogen, Camarillo, USA), anti-GAPDH (1:5000, #5174, Cell Signaling, Danvers, MA, USA) and anti-Actin (1:8000, #mab1501, Sigma–Aldrich, St. Louis, USA). The membrane was subsequently incubated with horseradish peroxidase–conjugated secondary antibodies, specifically goat anti-rabbit IgG (1:5000, Millipore, Billerica, USA) or goat anti-mouse IgG (1:5000, Millipore, Billerica, USA).). An Amersham Gel Imager (GE Healthcare, Chicago, Il, USA) and Quantity One software (Bio-Rad, Gladesville, NSW, Australia) were used for visualization and quantification of Western blot images. The quantitative results were normalized according to the corresponding GAPDH or ACTIN levels (as an internal control). Western blot analyses were performed on six randomly selected samples from each group, with each sample assayed in duplicate. Statistics Statistical analysis was conducted using SPSS software (V25.0, IBM, Armonk, NY, USA), while outliers were indentified and excluded using a Boxplot. The Kolmogorov-Smirnov test was used to assess data distribution. For normally distributed data, a two-way ANOVA (Exercise × Risperidone) was performed, followed by post-hoc least significant difference tests. For non-normally distributed data, a nonparametric Kruskal-Wallis H-test was performed, followed by post-hoc Mann–Whitney U-test was applied. Results are presented as the mean ± SEM, with p < 0.05 considered statistically significant. Results Hepatic lipid synthesis The risperidone-treated sedentary group showed increased protein expression of INSIG2, FAS and USF1, which was reduced by exercise intervention ( p < 0.05; Fig. 1 A, 1 B, 1 C). SCAP protein levels were increased in risperidone-treated groups (Fig. 1 D; p < 0.05). LXRα levels was decreased by risperidone treatment in the sedentary groups ( p < 0.05; Fig. 1 E). Voluntary exercise tended to increase the ratio of pAMPK/AMPK, while the co-treatment of risperidone and exercise further upregulated the ratio of pAMPK/AMPK ( p < 0.05; Fig. 1 F) significantly. There were no differences in precursor SREBP1c, mature SREBP1c, and its downstream target SCD1 and ACC1 (Fig. 1 G, 1 H, 1 I, 1 J). Hepatic lipid uptake and storage Hepatic levels of PPARγ and CD36 were increased by risperidone treatment and subsequently reversed by exercise intervention ( p < 0.05; Fig. 2 A, 2 B). FATP2 expression was reduced by exercise intervention in all groups treated with either vehicle or risperidone ( p < 0.05; Fig. 2 C). Reduced level of CAV-1 was observed in both the risperidone-only and exercise-only intervention groups (Fig. 2 D; p < 0.05). FSP27 expression was decreased by exercise intervention ( p < 0.05; Fig. 2 E). No significant difference was determined in FABP1 levels (Fig. 2 F). Hepatic lipolysis Although no significant difference was detected between the Risperidone + Sedentary and Vehicle + Sedentary groups, hepatic ATGL and HSL protein levels were higher in the Risperidone + Exercise than Risperidone + Sedentary groups ( p < 0.05; Fig. 3 A and 3 B). Hepatic fatty acid oxidation Reduced PGC1α expression was noted in the risperidone-only treatment group ( p < 0.05) that was reversed via exercise intervention (Fig. 3 C; p < 0.05). Discussion Our previous study found that exercise intervention reduces risperidone-induced elevations in plasma triglyceride levels, white adipose tissue weight, and insulin levels, suggesting altered lipid metabolism (Yi et al., 2021 ) The current study provides evidence that 4 weeks of voluntary exercise ameliorated risperidone-induced hepatic lipid metabolic disturbances by downregulating fatty acid synthesis (via USF1/FAS signalling) and uptake ( via PPARγ/CD36 signalling), while upregulating lipid breakdown ( via ATGL/HSL signalling) and fatty acid oxidation ( via PGC1α signalling) in female juvenile rats. AMPK regulates lipid metabolism by activating hepatic AMPK signaling, which inhibits the expression and activity of lipogenic regulators such as SREBP1—a critical transcription factor for de novo lipogenesis. Impairment of this regulatory axis contributes to lipid metabolism disorders. (Ferré and Foufelle, 2007 ). It is agreed with the reports that 4 weeks of risperidone treatment in this study did not change hepatic SREBP1c expression and activation of AMPK (Pozzi et al., 2024 ; Takami et al., 2010 ). It is noteworthy that the four-week exercise intervention increased the pAMPK/AMPK ratio in the risperidone treatment group. Exercise training is able to reduce triglyceride synthesis in muscle, white adipose tissue, and liver via the p-AMPK pathway (Kasper et al., 2021 ; Park et al., 2002 ). In this context, exercise intervention may confer benefits in decreasing triglyceride synthesis via the activation of the pAMPK pathway, although risperidone-enhanced lipid synthesis does not occur through this pathway. It has been reported that risperidone resulted in the overexpression of SREBP1c, SCAP, and its downstream lipogenic targets (SCD1, ACC1, and FAS), while downregulating INSIG2 (Auger et al., 2018 ; Cai et al., 2015 ). Only FAS expression was significantly upregulated by risperidone treatment in this study. It has been reported that risperidone could induce FAS without necessarily activating SREBP1c (Pozzi et al., 2024 ), while other transcription factors (e.g. LXR and USF1) may regulate FAS expression through both SREBP1-dependent and independent pathways (Griffin and Sul, 2004 ; Guo et al., 2018 ; Joseph et al., 2002 ). In fact, USF1 has been reported to mediate insulin-induced FAS expression (Griffin and Sul, 2004 ). USF1 and FAS expression levels were upregulated by risperidone treatment in this project, while this increase was reversed by exercise intervention, similar to the changes observed in plasma insulin levels (Sylvester et al., 2020 ). It suggests that exercise ameliorates risperidone-induced disturbances in lipogenesis through insulin/USF1/FAS signalling. Additionally, the risperidone-only treatment group exhibited increased INSIG2 expression and reduced LXRα expression, which does not match with previous findings (Auger et al., 2018 ; Cai et al., 2015 ). The discrepancy may be attributed to differences in animal gender, age, and treatment duration. Free fatty acids from blood are one of the primary sources of liver-derived fatty acids (Donnelly et al., 2005 ; Miles et al., 2004 ). Several proteins facilitate the influx of long-chain fatty acids into the liver, including scavenger receptor CD36, FATP2, CAV1, and FABP1 (Alves-Bezerra and Cohen, 2017 ). In addition, FSP27 enhances triglyceride accumulation (Xu et al., 2015 ). CD36 was found to be increased in animal models with hepatic steatosis, as well as patients with nonalcoholic fatty liver disease (Buqué et al., 2010 ; Miquilena-Colina et al., 2011 ). It remains unclear whether risperidone affects hepatic CD36 expression, while exercise intervention has been shown to suppress hepatic CD36 expression in mice with non-alcoholic steatohepatitis (Kawanishi et al., 2018 ). In addition, CD36 is a transcriptional target of PPARγ (Alves-Bezerra and Cohen, 2017 ; Tontonoz et al., 1998 ), which regulates liver triglyceride homeostasis (Gavrilova et al., 2003 ). We showed that risperidone increased levels of hepatic CD36 and PPARγ proteins, while exercise intervention decreased their levels. These results suggested that exercise could alleviate risperidone-induced hepatic lipometabolic disturbances through the PPARγ/CD36 pathway. Our study also observed that hepatic levels of PPARγ, CD36, and FAS proteins increased at 4 weeks, which aligns with findings from previous reports (Lee et al., 2018 ; Wang et al., 2020 ). CAV1, a major structural component of caveolae, plays a critical role in regulating hepatic lipid accumulation, glucose and lipid metabolism, and mitochondrial function. In our study, both risperidone and exercise interventions led to a reduction in hepatic CAV1 expression. Interestingly, previous studies have shown that CAV1-deficient mice are resistant to diet-induced obesity, while high-fat diet suppresses hepatic CAV1 expression (Deng et al., 2023 ; Razani et al., 2002 ). These findings collectively suggest that CAV1 plays a complex and context-dependent role in the regulation of lipid homeostasis. FATP2 is highly expressed in the liver, contributing to 40% of long-chain fatty acid uptake (Falcon et al., 2010 ). FABP1 facilitates the uptake, transport, and metabolism of fatty acids (Mashek, 2013 ). 12-week treatment of olanzapine has been reported to upregulate hepatic FATP2 and FABP1 expression (Jiang et al., 2019 ), whereas no difference was detected in the risperidone-treated groups in this study. Interestingly, exercise intervention reduced FATP2 and FSP27 levels. These results suggested that 4-weeks of voluntary exercise may reduce hepatic fatty acid uptake and lipid storage in juvenile rats. In addition to synthesis and uptake, fatty acids can be released from the hydrolysis of TGs, which is initiated by ATGL. ATGL and HSL are key enzymes in triacylglycerol catabolism, providing fatty acids (Brejchova et al., 2021 ) and promoting oxidation (Reid et al., 2008 ). It is agreed with our results that 8-week aerobic training has been reported to improve hepatic steatosis by promoting ATGL expression (Wu et al., 2022 ). In our study, 4 weeks of voluntary exercise increased hepatic ATGL and HSL protein expression in rats treated with risperidone, suggesting that ATGL and HSL may contribute to improved hepatic lipid metabolism through exercise. It is worth noting, however, that phosphorylated HSL (pHSL) and the pHSL/HSL ratio also play essential roles in lipolysis; future studies may benefit from evaluating these parameters. Most fatty acids from various sources will undergo mitochondrial β-oxidation to produce CO2 and ketone bodies (a main end product of hepatic FA catabolism) (Havel, 1972 ). PGC-1α enhances FA oxidation and reduces triacylglycerol storage and secretion in the liver (Morris et al., 2012 ). It has been documented that PGC-1α expression was downregulated by was decreased olanzapine in brown adipose tissue (Liu et al., 2020a ), while PGC-1α expression is increased by voluntary exercise in the liver (Rosa-Caldwell et al., 2017 ). Our findings demonstrated in the liver that voluntary exercise reversed risperidone-induced attenuation of PGC-1α, thereby improving lipid metabolism. PPARα, CPT1A and HMGCS2 play critical roles in hepatic fatty acid β-oxidation and ketogenesis (Dong et al., 2024 ; Houten et al., 2016 ; Kersten, 2014 ). Previous studies have indicated that both exercise and second-generation antipsychotics can affect their expression levels (Bae-Gartz et al., 2020 ; Chen et al., 2022 ). However, our previous study found that only PPARα expression was lower in the risperidone-only group (Yi et al., 2021 ). It is possible that four weeks of exercise intervention is insufficient to alter their expression levels. Conclusions In summary, chronic risperidone treatment in juvenile rats enhanced white adipose tissue accumulation, and upregulated fasting levels of triglyceride and insulin, which caused disturbances in lipid metabolism. However, a 4-week voluntary exercise intervention ameliorated these effects. This study revealed possible mechanisms: voluntary exercise intervention may decrease fatty acid synthesis through the insulin/USF1/FAS pathway, may reduce liver fatty acid uptake through the PPARγ/CD36 pathway, and may raise β-oxidation by up-regulating hepatic PGC1α expression, thereby improving risperidone-induced lipid disturbances. However, several limitations should be considered. Firstly, the lipid metabolism in drug-naïve patients with mental disorders is different from that in the healthy population (Penninx and Lange, 2018 ; Zhu et al., 2023 ). Therefore, an animal model for psychotic disorders will be valuable in future studies to investigate the mechanisms for risperidone and exercise interventions on lipid metabolism in patients with mental disorders. Secondly, the effects of voluntary exercise would be more pronounced if the running wheel were installed in their home cage, allowing the rats to access it at any time rather than just for 3 hours per day, as in this study. Overall, this project underscores the prospects of clinical exercise interventions in mitigating metabolic abnormalities in children/adolescents undergoing risperidone treatment. In addition to hepatic lipid regulation, white adipose tissue also plays a crucial role in lipid metabolism. Future studies will aim to investigate how risperidone and voluntary exercise modulate lipid metabolic pathways in adipose tissue. Furthermore, future research should explore the long-term benefits of lifelong exercise and its potential impacts during adolescence on adult health, particularly in juveniles with mental disorders. Declarations Funding Declaration This study was funded by the Australian National Health and Medical Research Council (NHMRC) Project Grant (APP1104184) to CD and JL. JL was also supported by an NHMRC Early Career Fellowship Award (APP1125937). The funding body did not play any roles in the design and conduct of the study, data interpretation and paper writing. Conflict of interest None of the authors has a conflict of interest. Author Contribution WY and CD designed the experiments. WY, and JL performed the experiments. WY and CD analyzed the data. WY prepared the initial draft of the manuscript. CD, WY, and JL revised the manuscript. All authors commented on and approved the final draft. Acknowledgement We thank Ms Emma Sylvester for her contributions in the animal experiment. Data Availability The datasets used during the current study are available from the corresponding author upon reasonable request. References Alsabhan JF et al. 2024. Metabolic Side Effects of Risperidone in Pediatric Patients with Neurological Disorders: A Prospective Cohort Study. J Clin Med. 13. Alves-Bezerra M, Cohen DE. Triglyceride Metabolism in the Liver. Compr Physiol. 2017;8:1–8. Auger F, et al. Risperidone-induced metabolic dysfunction is attenuated by Curcuma longa extract administration in mice. Metab Brain Dis. 2018;33:63–77. Bae-Gartz I, et al. Maternal exercise conveys protection against NAFLD in the offspring via hepatic metabolic programming. Sci Rep. 2020;10:15424. Brejchova K, et al. Distinct roles of adipose triglyceride lipase and hormone-sensitive lipase in the catabolism of triacylglycerol estolides. Proc Natl Acad Sci U S A; 2021. p. 118. Buqué X, et al. A subset of dysregulated metabolic and survival genes is associated with severity of hepatic steatosis in obese Zucker rats. J Lipid Res. 2010;51:500–13. Cai HL, et al. A potential mechanism underlying atypical antipsychotics-induced lipid disturbances. Transl Psychiatry. 2015;5:e661. Castellani LN et al. 2019. Preclinical and Clinical Sex Differences in Antipsychotic-Induced Metabolic Disturbances: A Narrative Review of Adiposity and Glucose Metabolism. J Psychiatr Brain Sci. 4. Chen CC, et al. Early Lipid Metabolic Effects of the Anti-Psychotic Drug Olanzapine on Weight Gain and the Associated Gene Expression. Neuropsychiatr Dis Treat. 2022;18:645–57. De Santis M, et al. Early antipsychotic treatment in childhood/adolescent period has long-term effects on depressive-like, anxiety-like and locomotor behaviours in adult rats. J Psychopharmacol. 2016;30:204–14. De Santis M, Huang XF, Deng C. Early antipsychotic treatment in juvenile rats elicits long-term alterations to the adult serotonin receptors. Neuropsychiatr Dis Treat. 2018;14:1569–83. Deng GH, et al. Caveolin-1 is critical for hepatic iron storage capacity in the development of nonalcoholic fatty liver disease. Mil Med Res. 2023;10:53. Dong J, et al. ACACA reduces lipid accumulation through dual regulation of lipid metabolism and mitochondrial function via AMPK- PPARα- CPT1A axis. J Transl Med. 2024;22:196. Donnelly KL, et al. Sources of fatty acids stored in liver and secreted via lipoproteins in patients with nonalcoholic fatty liver disease. J Clin Invest. 2005;115:1343–51. Falcon A, et al. FATP2 is a hepatic fatty acid transporter and peroxisomal very long-chain acyl-CoA synthetase. Am J Physiol Endocrinol Metab. 2010;299:E384–93. FDA. 2005. Guidance for Industry: Estimating the Maximum Safe Starting Dose in Initial Clinical Trials for Therapeutics in Adult Healthy Volunteers Vol., F.a.D. Administration, ed.^eds. Ferré P, Foufelle F. SREBP-1c transcription factor and lipid homeostasis: clinical perspective. Horm Res. 2007;68:72–82. Fitzgerald PB, et al. The relationship of changes in leptin, neuropeptide Y and reproductive hormones to antipsychotic induced weight gain. Hum Psychopharmacol. 2003;18:551–7. Gavrilova O, et al. Liver peroxisome proliferator-activated receptor gamma contributes to hepatic steatosis, triglyceride clearance, and regulation of body fat mass. J Biol Chem. 2003;278:34268–76. Gebhardt S, et al. Antipsychotic-induced body weight gain: predictors and a systematic categorization of the long-term weight course. J Psychiatr Res. 2009;43:620–6. Griffin MJ, Sul HS. Insulin regulation of fatty acid synthase gene transcription: roles of USF and SREBP-1c. IUBMB Life. 2004;56:595–600. Guo J, et al. Upstream stimulating factor 1 suppresses autophagy and hepatic lipid droplet catabolism by activating mTOR. FEBS Lett. 2018;592:2725–38. Havel RJ. Caloric homeostasis and disorders of fuel transport. N Engl J Med. 1972;287:1186–92. Houten SM, et al. The Biochemistry and Physiology of Mitochondrial Fatty Acid β-Oxidation and Its Genetic Disorders. Annu Rev Physiol. 2016;78:23–44. Jensen-Urstad AP, Semenkovich CF. Fatty acid synthase and liver triglyceride metabolism: housekeeper or messenger? Biochim Biophys Acta. 2012;1821:747–53. Jiang T, et al. Up-regulation of hepatic fatty acid transporters and inhibition/down-regulation of hepatic OCTN2 contribute to olanzapine-induced liver steatosis. Toxicol Lett. 2019;316:183–93. Joseph SB, et al. Direct and indirect mechanisms for regulation of fatty acid synthase gene expression by liver X receptors. J Biol Chem. 2002;277:11019–25. Kasper P et al. 2021. Maternal Exercise Mediates Hepatic Metabolic Programming via Activation of AMPK-PGC1α Axis in the Offspring of Obese Mothers. Cells. 10. Kawanishi N et al. 2018. Exercise training suppresses scavenger receptor CD36 expression in kupffer cells of nonalcoholic steatohepatitis model mice. Physiol Rep 6, e13902. Kelly DL, Conley RR, Tamminga CA. Differential olanzapine plasma concentrations by sex in a fixed-dose study. Schizophr Res. 1999;40:101–4. Kersten S. Integrated physiology and systems biology of PPARα. Mol Metab. 2014;3:354–71. Klau J et al. 2024. Antipsychotic prescribing patterns in children and adolescents attending Australian general practice in 2011 and 2017. JCPP Adv 4, e12208. Kurosaka Y et al. 2021. Protective Effects of Voluntary Exercise on Hepatic Fat Accumulation Induced by Dietary Restriction in Zucker Fatty Rats. Int J Mol Sci 22. Lee YK, et al. Hepatic lipid homeostasis by peroxisome proliferator-activated receptor gamma 2. Liver Res. 2018;2:209–15. Lian J, et al. Risperidone-induced weight gain and reduced locomotor activity in juvenile female rats: The role of histaminergic and NPY pathways. Pharmacol Res. 2015;95–96:20–6. Liu X, et al. Brown adipose tissue activity is modulated in olanzapine-treated young rats by simvastatin. BMC Pharmacol Toxicol. 2020a;21:48. Liu Y, et al. Hepatic Slug epigenetically promotes liver lipogenesis, fatty liver disease, and type 2 diabetes. J Clin Invest. 2020b;130:2992–3004. Mashek DG. Hepatic fatty acid trafficking: multiple forks in the road. Adv Nutr. 2013;4:697–710. May FJ, et al. Lipidomic Adaptations in White and Brown Adipose Tissue in Response to Exercise Demonstrate Molecular Species-Specific Remodeling. Cell Rep. 2017;18:1558–72. Miles JM, et al. Systemic and forearm triglyceride metabolism: fate of lipoprotein lipase-generated glycerol and free fatty acids. Diabetes. 2004;53:521–7. Miquilena-Colina ME, et al. Hepatic fatty acid translocase CD36 upregulation is associated with insulin resistance, hyperinsulinaemia and increased steatosis in non-alcoholic steatohepatitis and chronic hepatitis C. Gut. 2011;60:1394–402. Morrato EH, et al. Metabolic Screening in Children Receiving Antipsychotic Drug Treatment. Arch Pediatr Adolesc Med. 2010;164:344–51. Morris EM, et al. PGC-1α overexpression results in increased hepatic fatty acid oxidation with reduced triacylglycerol accumulation and secretion. Am J Physiol Gastrointest Liver Physiol. 2012;303:G979–92. Oh KJ, et al. Atypical antipsychotic drugs perturb AMPK-dependent regulation of hepatic lipid metabolism. Am J Physiol Endocrinol Metab. 2011;300:E624–32. Park H, et al. Coordinate regulation of malonyl-CoA decarboxylase, sn-glycerol-3-phosphate acyltransferase, and acetyl-CoA carboxylase by AMP-activated protein kinase in rat tissues in response to exercise. J Biol Chem. 2002;277:32571–7. Penninx B, Lange SMM. Metabolic syndrome in psychiatric patients: overview, mechanisms, and implications. Dialogues Clin Neurosci. 2018;20:63–73. Pillinger T, et al. Comparative effects of 18 antipsychotics on metabolic function in patients with schizophrenia, predictors of metabolic dysregulation, and association with psychopathology: a systematic review and network meta-analysis. Lancet Psychiatry. 2020;7:64–77. Pozzi M, et al. Olanzapine, risperidone and ziprasidone differently affect lysosomal function and autophagy, reflecting their different metabolic risk in patients. Transl Psychiatry. 2024;14:13. Razani B, et al. Caveolin-1-deficient Mice Are Lean, Resistant to Diet-induced Obesity, and Show Hypertriglyceridemia with Adipocyte Abnormalities*. J Biol Chem. 2002;277:8635–47. Reagan-Shaw S, Nihal M, Ahmad N. Dose translation from animal to human studies revisited. FASEB J. 2008;22:659–61. Reid BN, et al. Hepatic overexpression of hormone-sensitive lipase and adipose triglyceride lipase promotes fatty acid oxidation, stimulates direct release of free fatty acids, and ameliorates steatosis. J Biol Chem. 2008;283:13087–99. Rosa-Caldwell ME, et al. Moderate physical activity promotes basal hepatic autophagy in diet-induced obese mice. Appl Physiol Nutr Metab. 2017;42:148–56. Su Y, et al. Epigenetic histone modulations of PPARγ and related pathways contribute to olanzapine-induced metabolic disorders. Pharmacol Res. 2020;155:104703. Su Y et al. 2023. Epigenetic Histone Methylation of PPARγ and CPT1A Signaling Contributes to Betahistine Preventing Olanzapine-Induced Dyslipidemia. Int J Mol Sci 24. Sylvester E, et al. Exercise intervention for preventing risperidone-induced dyslipidemia and gluco-metabolic disorders in female juvenile rats. Pharmacol Biochem Behav. 2020;199:173064. Takami G, et al. Effects of atypical antipsychotics and haloperidol on PC12 cells: only aripiprazole phosphorylates AMP-activated protein kinase. J Neural Transm (Vienna). 2010;117:1139–53. Tontonoz P, et al. PPARgamma promotes monocyte/macrophage differentiation and uptake of oxidized LDL. Cell. 1998;93:241–52. Vega RB, Huss JM, Kelly DP. The coactivator PGC-1 cooperates with peroxisome proliferator-activated receptor alpha in transcriptional control of nuclear genes encoding mitochondrial fatty acid oxidation enzymes. Mol Cell Biol. 2000;20:1868–76. Wang Y et al. 2020. PPARs as Metabolic Regulators in the Liver: Lessons from Liver-Specific PPAR-Null Mice. Int J Mol Sci 21. Wooten JS, et al. The effects of voluntary wheel running during weight-loss on biomarkers of hepatic lipid metabolism and inflammation in C57Bl/6J mice. Curr Res Physiol. 2022;5:63–72. Wu B, et al. Aerobic exercise promotes the expression of ATGL and attenuates inflammation to improve hepatic steatosis via lncRNA SRA. Sci Rep. 2022;12:5370. Xu X, et al. Transcriptional activation of Fsp27 by the liver-enriched transcription factor CREBH promotes lipid droplet growth and hepatic steatosis. Hepatology. 2015;61:857–69. Yi W, et al. Kidney plays an important role in ketogenesis induced by risperidone and voluntary exercise in juvenile female rats. Psychiatry Res. 2021;305:114196. Zhu Q, et al. Prevalence and clinical correlates of abnormal lipid metabolism in first-episode and drug-naïve patients with major depressive disorder with abnormal glucose metabolism. Sci Rep. 2023;13:8078. Additional Declarations No competing interests reported. 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Also discoverable on Platform About Our Team In Review Editorial Policies Advisory Board Help Center Resources Author Services Accessibility API Access RSS feed Manage Cookie Preferences © Research Square 2026 | ISSN 2693-5015 (online) Privacy Policy Terms of Service Do Not Sell My Personal Information {"props":{"pageProps":{"initialData":{"identity":"rs-7309963","acceptedTermsAndConditions":true,"allowDirectSubmit":true,"archivedVersions":[],"articleType":"Research Article","associatedPublications":[],"authors":[{"id":502558038,"identity":"f3c116d7-a259-4f7f-9128-48b1c3ab22be","order_by":0,"name":"Weijie Yi","email":"","orcid":"","institution":"University of Wollongong","correspondingAuthor":false,"prefix":"","firstName":"Weijie","middleName":"","lastName":"Yi","suffix":""},{"id":502558039,"identity":"91dad3cc-3883-4fa3-9692-7b3174b735d7","order_by":1,"name":"Jiamei Lian","email":"","orcid":"","institution":"University of Wollongong","correspondingAuthor":false,"prefix":"","firstName":"Jiamei","middleName":"","lastName":"Lian","suffix":""},{"id":502558040,"identity":"2b0c1c1d-7e2b-483b-9ae6-c791db4048ce","order_by":2,"name":"Chao Deng","email":"data:image/png;base64,iVBORw0KGgoAAAANSUhEUgAAAZAAAAAyAQMAAABI0h/eAAAABlBMVEX///8AAABVwtN+AAAACXBIWXMAAA7EAAAOxAGVKw4bAAAArUlEQVRIiWNgGAWjYDACCSD+AGEaEK+FcQbJWph5SNLCP7v54WPbNpvEBvbmbRIMNYeJsOTOMWPj3La0xAaeY2USDMeI0GIgkWAmndt2OLdBIsdMgoGNKC3p36QtQVrk3wC1/CNKS46ZNCPYFh4zCSCDCL/cyCk27DmXVt/Gk1ZskdiXTlgL/4z0jQ9+lNkY87Mf3njjwzdrwlrggA1EJJCgYRSMglEwCkYBHgAAmTQzMYQa3EcAAAAASUVORK5CYII=","orcid":"","institution":"University of Wollongong","correspondingAuthor":true,"prefix":"","firstName":"Chao","middleName":"","lastName":"Deng","suffix":""}],"badges":[],"createdAt":"2025-08-06 12:53:19","currentVersionCode":1,"declarations":"","doi":"10.21203/rs.3.rs-7309963/v1","doiUrl":"https://doi.org/10.21203/rs.3.rs-7309963/v1","draftVersion":[],"editorialEvents":[],"editorialNote":"","failedWorkflow":false,"files":[{"id":89563393,"identity":"6571412e-c52d-44c8-9d4c-0d67b896af1d","added_by":"auto","created_at":"2025-08-21 10:30:14","extension":"png","order_by":1,"title":"Figure 1","display":"","copyAsset":false,"role":"figure","size":9557015,"visible":true,"origin":"","legend":"\u003cp\u003e\u003cstrong\u003eThe effects of risperidone and exercise intervention on the protein expression associated with hepatic lipogenesis. \u003c/strong\u003eWestern blot images and relative expression of (A) INSIG2, (B) FAS, (C) USF1, (D) SCAP, (E) LXRα, (F) pAMPk/AMPK, (G) p-SREBP1C, (H) m-SREBP1C, (I) SCD1, and (J) ACC1. Data represent Mean ± SEM (n=6/group). Abbreviations: VS, Vehicle+Sedentary group; RS, Risperidone+Sedentary group; VE, Vehicle+Exercise group; RE; Risperidone+Exercise group. *, \u003cem\u003ep\u003c/em\u003e\u0026lt;0.05; t, 0.05\u0026lt;\u003cem\u003ep\u003c/em\u003e\u0026lt;0.1 \u003cem\u003evs\u003c/em\u003e VS.\u003c/p\u003e","description":"","filename":"Fig1.png","url":"https://assets-eu.researchsquare.com/files/rs-7309963/v1/bb2526efd618a52bd7e05c2b.png"},{"id":89563394,"identity":"35e983da-6c2d-4156-a11c-4ffcdf50e695","added_by":"auto","created_at":"2025-08-21 10:30:14","extension":"png","order_by":2,"title":"Figure 2","display":"","copyAsset":false,"role":"figure","size":6194541,"visible":true,"origin":"","legend":"\u003cp\u003e\u003cstrong\u003eThe effects of risperidone and exercise intervention on the protein expression associated with fatty acid uptake. \u003c/strong\u003eWestern blot images and relative expression of (A) PPARγ, (B) CD36, (C) FATP2, (D) CAV-1, (E) FABP1, and (F) FSP27. Data represent Mean ± SEM (n = 6/group). Abbreviations: VS, Vehicle+Sedentary group; RS, Risperidone+Sedentary group; VE, Vehicle+Exercise group; RE; Risperidone+Exercise group. *, \u003cem\u003ep\u003c/em\u003e\u0026lt;0.05; t, 0.05\u0026lt;\u003cem\u003ep\u003c/em\u003e\u0026lt;0.1 \u003cem\u003evs \u003c/em\u003eVS.\u003c/p\u003e","description":"","filename":"Fig2.png","url":"https://assets-eu.researchsquare.com/files/rs-7309963/v1/9140651cd677a558e9c37f1d.png"},{"id":89563397,"identity":"8c6b3ce7-2714-4bfe-84e5-6dfcc7513120","added_by":"auto","created_at":"2025-08-21 10:30:14","extension":"png","order_by":3,"title":"Figure 3","display":"","copyAsset":false,"role":"figure","size":4183562,"visible":true,"origin":"","legend":"\u003cp\u003e\u003cstrong\u003eThe effect of risperidone and exercise intervention on the protein expression related to β-oxidation and lipolysis. \u003c/strong\u003eWestern blot images and the relative expression of (A) ATGL, (B) HSL, and (C) PGC1α. \u0026nbsp;Data represent Mean ± SEM (n = 6/group). Abbreviations: VS, Vehicle+Sedentary group; RS, Risperidone+Sedentary group; VE, Vehicle+Exercise group; RE; Risperidone+Exercise group. *, \u003cem\u003ep\u003c/em\u003e\u0026lt;0.05.\u003c/p\u003e","description":"","filename":"Fig3.png","url":"https://assets-eu.researchsquare.com/files/rs-7309963/v1/28417bee8aac1a2b559cc02e.png"},{"id":90351718,"identity":"8cebcd8d-08ce-47a1-85c0-b2bb57b48b04","added_by":"auto","created_at":"2025-09-01 18:01:41","extension":"pdf","order_by":0,"title":"","display":"","copyAsset":false,"role":"manuscript-pdf","size":19408707,"visible":true,"origin":"","legend":"","description":"","filename":"manuscript.pdf","url":"https://assets-eu.researchsquare.com/files/rs-7309963/v1/854a9ffd-6dd3-4860-888d-dec2f885258d.pdf"},{"id":89563388,"identity":"4d5fa858-de57-4cb1-babd-22a23f2c8dd6","added_by":"auto","created_at":"2025-08-21 10:30:14","extension":"docx","order_by":0,"title":"","display":"","copyAsset":false,"role":"supplement","size":18654,"visible":true,"origin":"","legend":"","description":"","filename":"SupplementaryTable1.docx","url":"https://assets-eu.researchsquare.com/files/rs-7309963/v1/1ac948a1d744cb81e6e8ca92.docx"},{"id":89563403,"identity":"2c3aab6a-6daa-4ec1-99b1-872735d96e9a","added_by":"auto","created_at":"2025-08-21 10:30:15","extension":"pdf","order_by":1,"title":"","display":"","copyAsset":false,"role":"supplement","size":2492179,"visible":true,"origin":"","legend":"","description":"","filename":"SupplementarymaterialsFullWesternblotsImages.pdf","url":"https://assets-eu.researchsquare.com/files/rs-7309963/v1/4ece17df543b8a4751984914.pdf"}],"financialInterests":"No competing interests reported.","formattedTitle":"The effect and mechanisms of risperidone and voluntary exercise intervention on hepatic lipid metabolism in juvenile female rats","fulltext":[{"header":"Introduction","content":"\u003cp\u003eRisperidone (neuroscience-based nomenclature (NbN): [dopamine D2, serotonin 5-HT2, and norepinephrine alpha-2 receptor antagonist]), one of the most commonly prescribed second-generation antipsychotics (SGAs), accounts for approximately 70% of antipsychotic prescriptions in juveniles under 14 years of age (Klau et al., \u003cspan citationid=\"CR32\" class=\"CitationRef\"\u003e2024\u003c/span\u003e). However, it is associated with significant metabolic side effects, including weight gain, insulin resistance, and dyslipidemia, which can lead to metabolic syndrome (Pillinger et al., \u003cspan citationid=\"CR47\" class=\"CitationRef\"\u003e2020\u003c/span\u003e). Vulnerable populations such as children, adolescents, and females are particularly susceptible to these adverse effects (Castellani et al., \u003cspan citationid=\"CR8\" class=\"CitationRef\"\u003e2019\u003c/span\u003e; Morrato et al., \u003cspan citationid=\"CR42\" class=\"CitationRef\"\u003e2010\u003c/span\u003e). A prospective study reported that more than half (53.8%) of pediatric patients receiving risperidone experienced at least one metabolic abnormality, with hyperlipidemia being the most common (34.6%)(Alsabhan et al., \u003cspan citationid=\"CR1\" class=\"CitationRef\"\u003e2024\u003c/span\u003e). Additionally, female sex, considered both a risk factor and a predictive marker for SGA-induced weight gain, likely reflects increased vulnerability due to sex-specific physiological characteristics, such as a higher proportion of adipose tissue and the modulatory influence of gonadal hormones(Fitzgerald et al., \u003cspan citationid=\"CR18\" class=\"CitationRef\"\u003e2003\u003c/span\u003e; Gebhardt et al., \u003cspan citationid=\"CR20\" class=\"CitationRef\"\u003e2009\u003c/span\u003e; Kelly et al., \u003cspan citationid=\"CR30\" class=\"CitationRef\"\u003e1999\u003c/span\u003e).\u003c/p\u003e\u003cp\u003eVoluntary exercise has been shown to improve lipid metabolism (Wooten et al., \u003cspan citationid=\"CR60\" class=\"CitationRef\"\u003e2022\u003c/span\u003e). In our previous work, we found that voluntary exercise significantly attenuated risperidone-induced increases in plasma triglyceride levels and adipose tissue accumulation in juvenile rats (Yi et al., \u003cspan citationid=\"CR63\" class=\"CitationRef\"\u003e2021\u003c/span\u003e). However, the underlying mechanisms responsible for these protective effects remain incompletely understood.\u003c/p\u003e\u003cp\u003eThe liver is essential in maintaining whole-body lipid homeostasis by regulating the synthesis, storage, modification, and transport of lipids. Hepatic \u003cem\u003ede novo\u003c/em\u003e lipogenesis contributes to the storage and secretion of lipids from hepatocytes (Jensen-Urstad and Semenkovich, \u003cspan citationid=\"CR25\" class=\"CitationRef\"\u003e2012\u003c/span\u003e). Insulin activates lipogenic transcription factors [sterol regulatory element binding transcription factor 1c (SREBP1c), liver X receptor (LXR), and upstream transcription factor 1 (USF1)] upregulate the expression of lipogenic enzymes [e.g. Fatty acid synthase (FAS), Acetyl-CoA carboxylase1 (ACC1) and Stearoyl-CoA desaturase (SCD1)], resulting in fatty acid synthesis (Liu et al., \u003cspan citationid=\"CR37\" class=\"CitationRef\"\u003e2020b\u003c/span\u003e). Synthesized fatty acids are stored in the liver on the form of triglycerides and exported into the bloodstream in very-low-density lipoprotein (VLDL) particles. Additionally, fatty acids could be released from triglycerides through the catalysis of adipose triglyceride lipase (ATGL) and hormone-sensitive lipase (HSL), and then plasma fatty acids are taken up into the liver by fatty acid transport protein 2 (FATP2), caveolin-1 (CAV1) and cluster of differentiation 36 (CD36). Moreover, peroxisome proliferator-activated receptor α (PPARα) and γ (PPARγ) also modulate fatty acid uptake, trafficking, catabolism, utilization, triglyceride synthesis, and lipid droplet formation (Dong et al., \u003cspan citationid=\"CR13\" class=\"CitationRef\"\u003e2024\u003c/span\u003e; Su et al., \u003cspan citationid=\"CR53\" class=\"CitationRef\"\u003e2020\u003c/span\u003e). Further, the majority of the fatty acids in hepatocytes is translocated into the mitochondria and undergo β-oxidation (Alves-Bezerra and Cohen, \u003cspan citationid=\"CR2\" class=\"CitationRef\"\u003e2017\u003c/span\u003e). Carnitine palmitoyltransferase 1A (CPT1A), a downstream target of PPARα and a rate-limiting enzyme for fatty acid β-oxidation, facilitating fatty acids entering the mitochondrial matrix (Houten et al., \u003cspan citationid=\"CR24\" class=\"CitationRef\"\u003e2016\u003c/span\u003e). Peroxisome proliferator-activated receptor-γ coactivator 1-α (PGC-1α) collaborates with PPARα to regulate the expression of fatty acid oxidation enzymes in mitochondria (Vega et al., \u003cspan citationid=\"CR58\" class=\"CitationRef\"\u003e2000\u003c/span\u003e). Imbalances between lipid synthesis and degradation leads to lipid metabolism disorders.\u003c/p\u003e\u003cp\u003eTreatment with SGAs has been reported to disrupt hepatic lipogenesis, lipolysis, fatty acids uptake, and β-oxidation (Oh et al., \u003cspan citationid=\"CR44\" class=\"CitationRef\"\u003e2011\u003c/span\u003e; Su et al., \u003cspan citationid=\"CR54\" class=\"CitationRef\"\u003e2023\u003c/span\u003e). Meanwhile, exercise improves lipid homeostasis by reducing synthesis and transport of fatty acids triglyceride in both adipose tissue and liver (Kurosaka et al., \u003cspan citationid=\"CR33\" class=\"CitationRef\"\u003e2021\u003c/span\u003e; May et al., \u003cspan citationid=\"CR39\" class=\"CitationRef\"\u003e2017\u003c/span\u003e). To date, no study has investigated the mechanisms through which exercise ameliorates lipid metabolism disorders induced by risperidone. Therefore, this study explored the possible mechanisms driving the effects of voluntary exercise in alleviating risperidone-induced lipid metabolic disorders in a juvenile female rat model.\u003c/p\u003e"},{"header":"Materials and Methods","content":"\u003cdiv id=\"Sec3\" class=\"Section2\"\u003e\u003ch2\u003eAnimal housing and treatment\u003c/h2\u003e\u003cp\u003eAnimal housing and treatment protocols were conducted as previously detailed (Yi et al., \u003cspan citationid=\"CR63\" class=\"CitationRef\"\u003e2021\u003c/span\u003e). Briefly, juvenile female Sprague-Dawley rats (postnatal day 22/23) were obtained from the Animal Resource Centre (Perth, Western Australia). At postnatal day 26/27, they were housed individually in Techniplast GR1800 ventilated cage (Lane Cove West, NSW, Australia), and randomly allocated into (1) Vehicle\u0026thinsp;+\u0026thinsp;Sedentary (VS), (2) Risperidone\u0026thinsp;+\u0026thinsp;Sedentary (RS), (3) Vehicle\u0026thinsp;+\u0026thinsp;Exercise (VE), and (4) Risperidone\u0026thinsp;+\u0026thinsp;Exercise (RS) groups (n\u0026thinsp;=\u0026thinsp;8/group). Risperidone (0.9 mg/kg; Janssen, Macquarie Park, NSW, Australia)was administered at a total dose of 1.8 mg/kg/day (0.9 mg/kg per dose, twice daily at 07:00 and 19:00) in 0.3 g cookie dough pellets from postnatal days 29/30 for a duration of 4 weeks, while the control rats were given plain cookie dough pellets (15% gelatine, 9% milk powder, 38% corn flour and 38% sugar) at the same times. The dosage was translated from the clinical dose based on body surface area in accordance with FDA guidelines (FDA, \u003cspan citationid=\"CR16\" class=\"CitationRef\"\u003e2005\u003c/span\u003e; Reagan-Shaw et al., \u003cspan citationid=\"CR50\" class=\"CitationRef\"\u003e2008\u003c/span\u003e), and has been shown to be physiologically and behaviorally effective in juvenile rats (De Santis et al., \u003cspan citationid=\"CR10\" class=\"CitationRef\"\u003e2016\u003c/span\u003e; De Santis et al., \u003cspan citationid=\"CR11\" class=\"CitationRef\"\u003e2018\u003c/span\u003e; Lian et al., \u003cspan citationid=\"CR35\" class=\"CitationRef\"\u003e2015\u003c/span\u003e; Sylvester et al., \u003cspan citationid=\"CR55\" class=\"CitationRef\"\u003e2020\u003c/span\u003e). On postnatal day 57/58, following overnight fasting, the final dose of risperidone was administered orally using a 1 mL syringe, with the drug dissolved in approximately 0.2 mL of water to avoid the potential impact of cookie dough on plasma glucose and lipid levels. Rats were allowed to voluntarily access running wheels equipped with revolution counters for 3 hours daily in a 4-week period (from postnatal days 29/30 to 56/57), with traveling distance recorded (Scurry Rat Running Wheel/Chamber, Lafayette Instrument, IN, USA). All rats were euthanized by decapitation following isoflurane anesthesia on postnatal day 57/58. Tissue samples (liver, inguinal, perirenal, periovary, and mesentery adipose tissue) were harvested and weighed immediately, and then frozen in liquid nitrogen and kept at -80\u0026deg;C. Blood was collected from the left ventricle into EDTA tube, and the plasma was separated by centrifuge (4℃, 3000 rpm, 10 min) then stored at -80\u0026deg;C until further use.\u003c/p\u003e\u003cp\u003eAs previously reported, Risperidone treatment significantly reduced physical activity over the 28-day intervention period (Average distance travelled: RE group, 1656.13\u0026thinsp;\u0026plusmn;\u0026thinsp;359.03 m/day vs VE group, 2828.00\u0026thinsp;\u0026plusmn;\u0026thinsp;416.24 m/day, p\u0026thinsp;\u0026lt;\u0026thinsp;0.05) (Yi et al., \u003cspan citationid=\"CR63\" class=\"CitationRef\"\u003e2021\u003c/span\u003e). Voluntary exercise reduced risperidone-induced increases in adipose tissue [periovary index (VS: 0.89\u0026thinsp;\u0026plusmn;\u0026thinsp;0.06, RS: 1.27\u0026thinsp;\u0026plusmn;\u0026thinsp;0.12, VE: 0.75\u0026thinsp;\u0026plusmn;\u0026thinsp;0.06, RE: 0.85\u0026thinsp;\u0026plusmn;\u0026thinsp;0.13), perirenal index (VS: 0.84\u0026thinsp;\u0026plusmn;\u0026thinsp;0.09, RS: 1.06\u0026thinsp;\u0026plusmn;\u0026thinsp;0.10, VE: 0.63\u0026thinsp;\u0026plusmn;\u0026thinsp;0.04, RE: 0.78\u0026thinsp;\u0026plusmn;\u0026thinsp;0.10) and inguinal (VS: 1.15\u0026thinsp;\u0026plusmn;\u0026thinsp;0.09, RS: 1.56\u0026thinsp;\u0026plusmn;\u0026thinsp;0.08, VE: 1.01\u0026thinsp;\u0026plusmn;\u0026thinsp;0.05, RE: 1.25\u0026thinsp;\u0026plusmn;\u0026thinsp;0.11)], fasting plasma insulin (VS: 85.27\u0026thinsp;\u0026plusmn;\u0026thinsp;6.35, RS: 201.16\u0026thinsp;\u0026plusmn;\u0026thinsp;48.08, VE: 129.27\u0026thinsp;\u0026plusmn;\u0026thinsp;19.37, RE: 113.67\u0026thinsp;\u0026plusmn;\u0026thinsp;16.76 pmol/L), and triglycerides(VS: 0.67\u0026thinsp;\u0026plusmn;\u0026thinsp;0.09, RS: 1.22\u0026thinsp;\u0026plusmn;\u0026thinsp;0.18, VE: 0.56\u0026thinsp;\u0026plusmn;\u0026thinsp;0.06, RE: 0.71\u0026thinsp;\u0026plusmn;\u0026thinsp;0.08 mM) (Yi et al., \u003cspan citationid=\"CR63\" class=\"CitationRef\"\u003e2021\u003c/span\u003e). The Animal Ethics Committee, University of Wollongong, Australia, approved all experimental procedures (AE18/19).\u003c/p\u003e\u003c/div\u003e\n\u003ch3\u003eWestern blots\u003c/h3\u003e\n\u003cp\u003eProcedures of the liver lysate preparations and Western blot were conducted as reported previously (Yi et al., \u003cspan citationid=\"CR63\" class=\"CitationRef\"\u003e2021\u003c/span\u003e). In brief, aliquots containing 15 \u0026micro;g protein were added to electrophoresis on a precast polyacrylamide (4\u0026ndash;20%) gel (Bio-Rad Laboratories, Gladesville, NSW, Australia). transferred the separated protein to a polyvinylidene difluoride (PVDF) membrane (Bio-Rad Laboratories, Gladesville, NSW, Australia). Following transferred the separated protein to a polyvinylidene difluoride membrane, it was blocked with 5% skim milk plus 0.1% Tween-20 in Tris-buffered saline for a subsequent overnight incubation at 4\u0026deg;C with following primary antibodies: anti-SREBP1 (1:500, #ab28481, Abcam, Cambridge, UK), anti-SCD1 (1:1000, #ab19862, Abcam, Cambridge, UK), anti-PPARγ (1:1000, ab209350, Abcam, Cambridge, UK), anti-USF1 (1:1000, #ab180717, Abcam, Cambridge, UK), anti-ATGL (1:1000, #ab109251, Abcam, Cambridge, UK), anti-FSP27 (1:1000, #ab213693, Abcam, Cambridge, UK), anti-PGC1α (1:1000, #ab191838, Abcam, Cambridge, UK), anti-LXRα(1:1000, #ab106464, Abcam, Cambridge, UK), CAV-1(1:1000, #ab2910, Abcam, Cambridge, UK) HSL(1:1000, #ab45422, Abcam, Cambridge, UK), FABP1 (1:1000, #ab222517, Abcam, Cambridge, UK), anti-FAS (1:1000, #3180S, Cell Signaling, Danvers, MA, USA), anti-CD36 (1:1000, #74002, Cell Signaling, Danvers, MA, USA), anti-SCAP (1:1000, #13102S, Cell Signaling, Danvers, MA, USA), anti-pAMPKα(1:2000, #2535S, Cell Signaling, Danvers, MA, USA), anti-AMPKα(1:1000, #2532, Cell Signaling, Danvers, MA, USA), anti-ACC (1:500, #3662S, Cell Signaling, Danvers, MA, USA) anti-INSIG2 (1:1000, #PA5109863, Invitrogen, Camarillo, USA), anti-FATP2 (1:1000, #MA5-50447, Invitrogen, Camarillo, USA), anti-GAPDH (1:5000, #5174, Cell Signaling, Danvers, MA, USA) and anti-Actin (1:8000, #mab1501, Sigma\u0026ndash;Aldrich, St. Louis, USA). The membrane was subsequently incubated with horseradish peroxidase\u0026ndash;conjugated secondary antibodies, specifically goat anti-rabbit IgG (1:5000, Millipore, Billerica, USA) or goat anti-mouse IgG (1:5000, Millipore, Billerica, USA).). An Amersham Gel Imager (GE Healthcare, Chicago, Il, USA) and Quantity One software (Bio-Rad, Gladesville, NSW, Australia) were used for visualization and quantification of Western blot images. The quantitative results were normalized according to the corresponding GAPDH or ACTIN levels (as an internal control). Western blot analyses were performed on six randomly selected samples from each group, with each sample assayed in duplicate.\u003c/p\u003e\n\u003ch3\u003eStatistics\u003c/h3\u003e\n\u003cp\u003eStatistical analysis was conducted using SPSS software (V25.0, IBM, Armonk, NY, USA), while outliers were indentified and excluded using a Boxplot. The Kolmogorov-Smirnov test was used to assess data distribution. For normally distributed data, a two-way ANOVA (Exercise \u0026times; Risperidone) was performed, followed by post-hoc least significant difference tests. For non-normally distributed data, a nonparametric Kruskal-Wallis H-test was performed, followed by post-hoc Mann\u0026ndash;Whitney U-test was applied. Results are presented as the mean\u0026thinsp;\u0026plusmn;\u0026thinsp;SEM, with \u003cem\u003ep\u003c/em\u003e\u0026thinsp;\u0026lt;\u0026thinsp;0.05 considered statistically significant.\u003c/p\u003e"},{"header":"Results","content":"\u003cdiv id=\"Sec7\" class=\"Section2\"\u003e\u003ch2\u003eHepatic lipid synthesis\u003c/h2\u003e\u003cp\u003eThe risperidone-treated sedentary group showed increased protein expression of INSIG2, FAS and USF1, which was reduced by exercise intervention (\u003cem\u003ep\u003c/em\u003e\u0026thinsp;\u0026lt;\u0026thinsp;0.05; Fig.\u0026nbsp;\u003cspan refid=\"Fig1\" class=\"InternalRef\"\u003e1\u003c/span\u003eA, \u003cspan refid=\"Fig1\" class=\"InternalRef\"\u003e1\u003c/span\u003eB, \u003cspan refid=\"Fig1\" class=\"InternalRef\"\u003e1\u003c/span\u003eC). SCAP protein levels were increased in risperidone-treated groups (Fig.\u0026nbsp;\u003cspan refid=\"Fig1\" class=\"InternalRef\"\u003e1\u003c/span\u003eD; \u003cem\u003ep\u003c/em\u003e\u0026thinsp;\u0026lt;\u0026thinsp;0.05). LXRα levels was decreased by risperidone treatment in the sedentary groups (\u003cem\u003ep\u003c/em\u003e\u0026thinsp;\u0026lt;\u0026thinsp;0.05; Fig.\u0026nbsp;\u003cspan refid=\"Fig1\" class=\"InternalRef\"\u003e1\u003c/span\u003eE). Voluntary exercise tended to increase the ratio of pAMPK/AMPK, while the co-treatment of risperidone and exercise further upregulated the ratio of pAMPK/AMPK (\u003cem\u003ep\u003c/em\u003e\u0026thinsp;\u0026lt;\u0026thinsp;0.05; Fig.\u0026nbsp;\u003cspan refid=\"Fig1\" class=\"InternalRef\"\u003e1\u003c/span\u003eF) significantly. There were no differences in precursor SREBP1c, mature SREBP1c, and its downstream target SCD1 and ACC1 (Fig.\u0026nbsp;\u003cspan refid=\"Fig1\" class=\"InternalRef\"\u003e1\u003c/span\u003eG, \u003cspan refid=\"Fig1\" class=\"InternalRef\"\u003e1\u003c/span\u003eH, \u003cspan refid=\"Fig1\" class=\"InternalRef\"\u003e1\u003c/span\u003eI, \u003cspan refid=\"Fig1\" class=\"InternalRef\"\u003e1\u003c/span\u003eJ).\u003c/p\u003e\u003cp\u003e\u003c/p\u003e\u003c/div\u003e\u003cdiv id=\"Sec8\" class=\"Section2\"\u003e\u003ch2\u003eHepatic lipid uptake and storage\u003c/h2\u003e\u003cp\u003eHepatic levels of PPARγ and CD36 were increased by risperidone treatment and subsequently reversed by exercise intervention (\u003cem\u003ep\u003c/em\u003e\u0026thinsp;\u0026lt;\u0026thinsp;0.05; Fig.\u0026nbsp;\u003cspan refid=\"Fig2\" class=\"InternalRef\"\u003e2\u003c/span\u003eA, \u003cspan refid=\"Fig2\" class=\"InternalRef\"\u003e2\u003c/span\u003eB). FATP2 expression was reduced by exercise intervention in all groups treated with either vehicle or risperidone (\u003cem\u003ep\u003c/em\u003e\u0026thinsp;\u0026lt;\u0026thinsp;0.05; Fig.\u0026nbsp;\u003cspan refid=\"Fig2\" class=\"InternalRef\"\u003e2\u003c/span\u003eC). Reduced level of CAV-1 was observed in both the risperidone-only and exercise-only intervention groups (Fig.\u0026nbsp;\u003cspan refid=\"Fig2\" class=\"InternalRef\"\u003e2\u003c/span\u003eD; \u003cem\u003ep\u003c/em\u003e\u0026thinsp;\u0026lt;\u0026thinsp;0.05). FSP27 expression was decreased by exercise intervention (\u003cem\u003ep\u003c/em\u003e\u0026thinsp;\u0026lt;\u0026thinsp;0.05; Fig.\u0026nbsp;\u003cspan refid=\"Fig2\" class=\"InternalRef\"\u003e2\u003c/span\u003eE). No significant difference was determined in FABP1 levels (Fig.\u0026nbsp;\u003cspan refid=\"Fig2\" class=\"InternalRef\"\u003e2\u003c/span\u003eF).\u003c/p\u003e\u003cp\u003e\u003c/p\u003e\u003c/div\u003e\n\u003ch3\u003eHepatic lipolysis\u003c/h3\u003e\n\u003cp\u003eAlthough no significant difference was detected between the Risperidone\u0026thinsp;+\u0026thinsp;Sedentary and Vehicle\u0026thinsp;+\u0026thinsp;Sedentary groups, hepatic ATGL and HSL protein levels were higher in the Risperidone\u0026thinsp;+\u0026thinsp;Exercise than Risperidone\u0026thinsp;+\u0026thinsp;Sedentary groups (\u003cem\u003ep\u003c/em\u003e\u0026thinsp;\u0026lt;\u0026thinsp;0.05; Fig.\u0026nbsp;\u003cspan refid=\"Fig3\" class=\"InternalRef\"\u003e3\u003c/span\u003eA and \u003cspan refid=\"Fig3\" class=\"InternalRef\"\u003e3\u003c/span\u003eB).\u003c/p\u003e\u003cp\u003e\u003c/p\u003e\n\u003ch3\u003eHepatic fatty acid oxidation\u003c/h3\u003e\n\u003cp\u003eReduced PGC1α expression was noted in the risperidone-only treatment group (\u003cem\u003ep\u003c/em\u003e\u0026thinsp;\u0026lt;\u0026thinsp;0.05) that was reversed via exercise intervention (Fig.\u0026nbsp;\u003cspan refid=\"Fig3\" class=\"InternalRef\"\u003e3\u003c/span\u003eC; \u003cem\u003ep\u003c/em\u003e\u0026thinsp;\u0026lt;\u0026thinsp;0.05).\u003c/p\u003e"},{"header":"Discussion","content":"\u003cp\u003eOur previous study found that exercise intervention reduces risperidone-induced elevations in plasma triglyceride levels, white adipose tissue weight, and insulin levels, suggesting altered lipid metabolism (Yi et al., \u003cspan citationid=\"CR63\" class=\"CitationRef\"\u003e2021\u003c/span\u003e) The current study provides evidence that 4 weeks of voluntary exercise ameliorated risperidone-induced hepatic lipid metabolic disturbances by downregulating fatty acid synthesis (via USF1/FAS signalling) and uptake (\u003cem\u003evia\u003c/em\u003e PPARγ/CD36 signalling), while upregulating lipid breakdown (\u003cem\u003evia\u003c/em\u003e ATGL/HSL signalling) and fatty acid oxidation (\u003cem\u003evia\u003c/em\u003e PGC1α signalling) in female juvenile rats.\u003c/p\u003e\u003cp\u003eAMPK regulates lipid metabolism by activating hepatic AMPK signaling, which inhibits the expression and activity of lipogenic regulators such as SREBP1\u0026mdash;a critical transcription factor for de novo lipogenesis. Impairment of this regulatory axis contributes to lipid metabolism disorders. (Ferr\u0026eacute; and Foufelle, \u003cspan citationid=\"CR17\" class=\"CitationRef\"\u003e2007\u003c/span\u003e). It is agreed with the reports that 4 weeks of risperidone treatment in this study did not change hepatic SREBP1c expression and activation of AMPK (Pozzi et al., \u003cspan citationid=\"CR48\" class=\"CitationRef\"\u003e2024\u003c/span\u003e; Takami et al., \u003cspan citationid=\"CR56\" class=\"CitationRef\"\u003e2010\u003c/span\u003e). It is noteworthy that the four-week exercise intervention increased the pAMPK/AMPK ratio in the risperidone treatment group. Exercise training is able to reduce triglyceride synthesis in muscle, white adipose tissue, and liver via the p-AMPK pathway (Kasper et al., \u003cspan citationid=\"CR28\" class=\"CitationRef\"\u003e2021\u003c/span\u003e; Park et al., \u003cspan citationid=\"CR45\" class=\"CitationRef\"\u003e2002\u003c/span\u003e). In this context, exercise intervention may confer benefits in decreasing triglyceride synthesis \u003cem\u003evia\u003c/em\u003e the activation of the pAMPK pathway, although risperidone-enhanced lipid synthesis does not occur through this pathway.\u003c/p\u003e\u003cp\u003eIt has been reported that risperidone resulted in the overexpression of SREBP1c, SCAP, and its downstream lipogenic targets (SCD1, ACC1, and FAS), while downregulating INSIG2 (Auger et al., \u003cspan citationid=\"CR3\" class=\"CitationRef\"\u003e2018\u003c/span\u003e; Cai et al., \u003cspan citationid=\"CR7\" class=\"CitationRef\"\u003e2015\u003c/span\u003e). Only FAS expression was significantly upregulated by risperidone treatment in this study. It has been reported that risperidone could induce FAS without necessarily activating SREBP1c (Pozzi et al., \u003cspan citationid=\"CR48\" class=\"CitationRef\"\u003e2024\u003c/span\u003e), while other transcription factors (e.g. LXR and USF1) may regulate FAS expression through both SREBP1-dependent and independent pathways (Griffin and Sul, \u003cspan citationid=\"CR21\" class=\"CitationRef\"\u003e2004\u003c/span\u003e; Guo et al., \u003cspan citationid=\"CR22\" class=\"CitationRef\"\u003e2018\u003c/span\u003e; Joseph et al., \u003cspan citationid=\"CR27\" class=\"CitationRef\"\u003e2002\u003c/span\u003e). In fact, USF1 has been reported to mediate insulin-induced FAS expression (Griffin and Sul, \u003cspan citationid=\"CR21\" class=\"CitationRef\"\u003e2004\u003c/span\u003e). USF1 and FAS expression levels were upregulated by risperidone treatment in this project, while this increase was reversed by exercise intervention, similar to the changes observed in plasma insulin levels (Sylvester et al., \u003cspan citationid=\"CR55\" class=\"CitationRef\"\u003e2020\u003c/span\u003e). It suggests that exercise ameliorates risperidone-induced disturbances in lipogenesis through insulin/USF1/FAS signalling. Additionally, the risperidone-only treatment group exhibited increased INSIG2 expression and reduced LXRα expression, which does not match with previous findings (Auger et al., \u003cspan citationid=\"CR3\" class=\"CitationRef\"\u003e2018\u003c/span\u003e; Cai et al., \u003cspan citationid=\"CR7\" class=\"CitationRef\"\u003e2015\u003c/span\u003e). The discrepancy may be attributed to differences in animal gender, age, and treatment duration.\u003c/p\u003e\u003cp\u003eFree fatty acids from blood are one of the primary sources of liver-derived fatty acids (Donnelly et al., \u003cspan citationid=\"CR14\" class=\"CitationRef\"\u003e2005\u003c/span\u003e; Miles et al., \u003cspan citationid=\"CR40\" class=\"CitationRef\"\u003e2004\u003c/span\u003e). Several proteins facilitate the influx of long-chain fatty acids into the liver, including scavenger receptor CD36, FATP2, CAV1, and FABP1 (Alves-Bezerra and Cohen, \u003cspan citationid=\"CR2\" class=\"CitationRef\"\u003e2017\u003c/span\u003e). In addition, FSP27 enhances triglyceride accumulation (Xu et al., \u003cspan citationid=\"CR62\" class=\"CitationRef\"\u003e2015\u003c/span\u003e). CD36 was found to be increased in animal models with hepatic steatosis, as well as patients with nonalcoholic fatty liver disease (Buqu\u0026eacute; et al., \u003cspan citationid=\"CR6\" class=\"CitationRef\"\u003e2010\u003c/span\u003e; Miquilena-Colina et al., \u003cspan citationid=\"CR41\" class=\"CitationRef\"\u003e2011\u003c/span\u003e). It remains unclear whether risperidone affects hepatic CD36 expression, while exercise intervention has been shown to suppress hepatic CD36 expression in mice with non-alcoholic steatohepatitis (Kawanishi et al., \u003cspan citationid=\"CR29\" class=\"CitationRef\"\u003e2018\u003c/span\u003e). In addition, CD36 is a transcriptional target of PPARγ (Alves-Bezerra and Cohen, \u003cspan citationid=\"CR2\" class=\"CitationRef\"\u003e2017\u003c/span\u003e; Tontonoz et al., \u003cspan citationid=\"CR57\" class=\"CitationRef\"\u003e1998\u003c/span\u003e), which regulates liver triglyceride homeostasis (Gavrilova et al., \u003cspan citationid=\"CR19\" class=\"CitationRef\"\u003e2003\u003c/span\u003e). We showed that risperidone increased levels of hepatic CD36 and PPARγ proteins, while exercise intervention decreased their levels. These results suggested that exercise could alleviate risperidone-induced hepatic lipometabolic disturbances through the PPARγ/CD36 pathway. Our study also observed that hepatic levels of PPARγ, CD36, and FAS proteins increased at 4 weeks, which aligns with findings from previous reports (Lee et al., \u003cspan citationid=\"CR34\" class=\"CitationRef\"\u003e2018\u003c/span\u003e; Wang et al., \u003cspan citationid=\"CR59\" class=\"CitationRef\"\u003e2020\u003c/span\u003e).\u003c/p\u003e\u003cp\u003eCAV1, a major structural component of caveolae, plays a critical role in regulating hepatic lipid accumulation, glucose and lipid metabolism, and mitochondrial function. In our study, both risperidone and exercise interventions led to a reduction in hepatic CAV1 expression. Interestingly, previous studies have shown that CAV1-deficient mice are resistant to diet-induced obesity, while high-fat diet suppresses hepatic CAV1 expression (Deng et al., \u003cspan citationid=\"CR12\" class=\"CitationRef\"\u003e2023\u003c/span\u003e; Razani et al., \u003cspan citationid=\"CR49\" class=\"CitationRef\"\u003e2002\u003c/span\u003e). These findings collectively suggest that CAV1 plays a complex and context-dependent role in the regulation of lipid homeostasis. FATP2 is highly expressed in the liver, contributing to 40% of long-chain fatty acid uptake (Falcon et al., \u003cspan citationid=\"CR15\" class=\"CitationRef\"\u003e2010\u003c/span\u003e). FABP1 facilitates the uptake, transport, and metabolism of fatty acids (Mashek, \u003cspan citationid=\"CR38\" class=\"CitationRef\"\u003e2013\u003c/span\u003e). 12-week treatment of olanzapine has been reported to upregulate hepatic FATP2 and FABP1 expression (Jiang et al., \u003cspan citationid=\"CR26\" class=\"CitationRef\"\u003e2019\u003c/span\u003e), whereas no difference was detected in the risperidone-treated groups in this study. Interestingly, exercise intervention reduced FATP2 and FSP27 levels. These results suggested that 4-weeks of voluntary exercise may reduce hepatic fatty acid uptake and lipid storage in juvenile rats.\u003c/p\u003e\u003cp\u003eIn addition to synthesis and uptake, fatty acids can be released from the hydrolysis of TGs, which is initiated by ATGL. ATGL and HSL are key enzymes in triacylglycerol catabolism, providing fatty acids (Brejchova et al., \u003cspan citationid=\"CR5\" class=\"CitationRef\"\u003e2021\u003c/span\u003e) and promoting oxidation (Reid et al., \u003cspan citationid=\"CR51\" class=\"CitationRef\"\u003e2008\u003c/span\u003e). It is agreed with our results that 8-week aerobic training has been reported to improve hepatic steatosis by promoting ATGL expression (Wu et al., \u003cspan citationid=\"CR61\" class=\"CitationRef\"\u003e2022\u003c/span\u003e). In our study, 4 weeks of voluntary exercise increased hepatic ATGL and HSL protein expression in rats treated with risperidone, suggesting that ATGL and HSL may contribute to improved hepatic lipid metabolism through exercise. It is worth noting, however, that phosphorylated HSL (pHSL) and the pHSL/HSL ratio also play essential roles in lipolysis; future studies may benefit from evaluating these parameters. Most fatty acids from various sources will undergo mitochondrial β-oxidation to produce CO2 and ketone bodies (a main end product of hepatic FA catabolism) (Havel, \u003cspan citationid=\"CR23\" class=\"CitationRef\"\u003e1972\u003c/span\u003e). PGC-1α enhances FA oxidation and reduces triacylglycerol storage and secretion in the liver (Morris et al., \u003cspan citationid=\"CR43\" class=\"CitationRef\"\u003e2012\u003c/span\u003e). It has been documented that PGC-1α expression was downregulated by was decreased olanzapine in brown adipose tissue (Liu et al., \u003cspan citationid=\"CR36\" class=\"CitationRef\"\u003e2020a\u003c/span\u003e), while PGC-1α expression is increased by voluntary exercise in the liver (Rosa-Caldwell et al., \u003cspan citationid=\"CR52\" class=\"CitationRef\"\u003e2017\u003c/span\u003e). Our findings demonstrated in the liver that voluntary exercise reversed risperidone-induced attenuation of PGC-1α, thereby improving lipid metabolism.\u003c/p\u003e\u003cp\u003ePPARα, CPT1A and HMGCS2 play critical roles in hepatic fatty acid β-oxidation and ketogenesis (Dong et al., \u003cspan citationid=\"CR13\" class=\"CitationRef\"\u003e2024\u003c/span\u003e; Houten et al., \u003cspan citationid=\"CR24\" class=\"CitationRef\"\u003e2016\u003c/span\u003e; Kersten, \u003cspan citationid=\"CR31\" class=\"CitationRef\"\u003e2014\u003c/span\u003e). Previous studies have indicated that both exercise and second-generation antipsychotics can affect their expression levels (Bae-Gartz et al., \u003cspan citationid=\"CR4\" class=\"CitationRef\"\u003e2020\u003c/span\u003e; Chen et al., \u003cspan citationid=\"CR9\" class=\"CitationRef\"\u003e2022\u003c/span\u003e). However, our previous study found that only PPARα expression was lower in the risperidone-only group (Yi et al., \u003cspan citationid=\"CR63\" class=\"CitationRef\"\u003e2021\u003c/span\u003e). It is possible that four weeks of exercise intervention is insufficient to alter their expression levels.\u003c/p\u003e"},{"header":"Conclusions","content":"\u003cp\u003eIn summary, chronic risperidone treatment in juvenile rats enhanced white adipose tissue accumulation, and upregulated fasting levels of triglyceride and insulin, which caused disturbances in lipid metabolism. However, a 4-week voluntary exercise intervention ameliorated these effects. This study revealed possible mechanisms: voluntary exercise intervention may decrease fatty acid synthesis through the insulin/USF1/FAS pathway, may reduce liver fatty acid uptake through the PPARγ/CD36 pathway, and may raise β-oxidation by up-regulating hepatic PGC1α expression, thereby improving risperidone-induced lipid disturbances. However, several limitations should be considered. Firstly, the lipid metabolism in drug-na\u0026iuml;ve patients with mental disorders is different from that in the healthy population (Penninx and Lange, \u003cspan citationid=\"CR46\" class=\"CitationRef\"\u003e2018\u003c/span\u003e; Zhu et al., \u003cspan citationid=\"CR64\" class=\"CitationRef\"\u003e2023\u003c/span\u003e). Therefore, an animal model for psychotic disorders will be valuable in future studies to investigate the mechanisms for risperidone and exercise interventions on lipid metabolism in patients with mental disorders. Secondly, the effects of voluntary exercise would be more pronounced if the running wheel were installed in their home cage, allowing the rats to access it at any time rather than just for 3 hours per day, as in this study. Overall, this project underscores the prospects of clinical exercise interventions in mitigating metabolic abnormalities in children/adolescents undergoing risperidone treatment. In addition to hepatic lipid regulation, white adipose tissue also plays a crucial role in lipid metabolism. Future studies will aim to investigate how risperidone and voluntary exercise modulate lipid metabolic pathways in adipose tissue. Furthermore, future research should explore the long-term benefits of lifelong exercise and its potential impacts during adolescence on adult health, particularly in juveniles with mental disorders.\u003c/p\u003e"},{"header":"Declarations","content":"\u003cp\u003e\u003cstrong\u003eFunding Declaration\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003eThis study was funded by the Australian National Health and Medical Research Council (NHMRC) Project Grant (APP1104184) to CD and JL. JL was also supported by an NHMRC Early Career Fellowship Award (APP1125937). The funding body did not play any roles in the design and conduct of the study, data interpretation and paper writing.\u003c/p\u003e\u003ch2\u003eConflict of interest\u003c/h2\u003e\u003cp\u003eNone of the authors has a conflict of interest.\u003c/p\u003e\u003c/p\u003e\u003ch2\u003eAuthor Contribution\u003c/h2\u003e\u003cp\u003eWY and CD designed the experiments. WY, and JL performed the experiments. WY and CD analyzed the data. WY prepared the initial draft of the manuscript. CD, WY, and JL revised the manuscript. All authors commented on and approved the final draft.\u003c/p\u003e\u003ch2\u003eAcknowledgement\u003c/h2\u003e\u003cp\u003eWe thank Ms Emma Sylvester for her contributions in the animal experiment.\u003c/p\u003e\u003ch2\u003eData Availability\u003c/h2\u003e\u003cp\u003eThe datasets used during the current study are available from the corresponding author upon reasonable request.\u003c/p\u003e"},{"header":"References","content":"\u003col\u003e\u003cli\u003e\u003cspan\u003eAlsabhan JF et al. 2024. Metabolic Side Effects of Risperidone in Pediatric Patients with Neurological Disorders: A Prospective Cohort Study. J Clin Med. 13.\u003c/span\u003e\u003c/li\u003e\u003cli\u003e\u003cspan\u003eAlves-Bezerra M, Cohen DE. Triglyceride Metabolism in the Liver. Compr Physiol. 2017;8:1\u0026ndash;8.\u003c/span\u003e\u003c/li\u003e\u003cli\u003e\u003cspan\u003eAuger F, et al. Risperidone-induced metabolic dysfunction is attenuated by Curcuma longa extract administration in mice. Metab Brain Dis. 2018;33:63\u0026ndash;77.\u003c/span\u003e\u003c/li\u003e\u003cli\u003e\u003cspan\u003eBae-Gartz I, et al. Maternal exercise conveys protection against NAFLD in the offspring via hepatic metabolic programming. Sci Rep. 2020;10:15424.\u003c/span\u003e\u003c/li\u003e\u003cli\u003e\u003cspan\u003eBrejchova K, et al. Distinct roles of adipose triglyceride lipase and hormone-sensitive lipase in the catabolism of triacylglycerol estolides. Proc Natl Acad Sci U S A; 2021. p. 118.\u003c/span\u003e\u003c/li\u003e\u003cli\u003e\u003cspan\u003eBuqu\u0026eacute; X, et al. A subset of dysregulated metabolic and survival genes is associated with severity of hepatic steatosis in obese Zucker rats. J Lipid Res. 2010;51:500\u0026ndash;13.\u003c/span\u003e\u003c/li\u003e\u003cli\u003e\u003cspan\u003eCai HL, et al. A potential mechanism underlying atypical antipsychotics-induced lipid disturbances. Transl Psychiatry. 2015;5:e661.\u003c/span\u003e\u003c/li\u003e\u003cli\u003e\u003cspan\u003eCastellani LN et al. 2019. Preclinical and Clinical Sex Differences in Antipsychotic-Induced Metabolic Disturbances: A Narrative Review of Adiposity and Glucose Metabolism. J Psychiatr Brain Sci. 4.\u003c/span\u003e\u003c/li\u003e\u003cli\u003e\u003cspan\u003eChen CC, et al. Early Lipid Metabolic Effects of the Anti-Psychotic Drug Olanzapine on Weight Gain and the Associated Gene Expression. Neuropsychiatr Dis Treat. 2022;18:645\u0026ndash;57.\u003c/span\u003e\u003c/li\u003e\u003cli\u003e\u003cspan\u003eDe Santis M, et al. Early antipsychotic treatment in childhood/adolescent period has long-term effects on depressive-like, anxiety-like and locomotor behaviours in adult rats. J Psychopharmacol. 2016;30:204\u0026ndash;14.\u003c/span\u003e\u003c/li\u003e\u003cli\u003e\u003cspan\u003eDe Santis M, Huang XF, Deng C. Early antipsychotic treatment in juvenile rats elicits long-term alterations to the adult serotonin receptors. Neuropsychiatr Dis Treat. 2018;14:1569\u0026ndash;83.\u003c/span\u003e\u003c/li\u003e\u003cli\u003e\u003cspan\u003eDeng GH, et al. Caveolin-1 is critical for hepatic iron storage capacity in the development of nonalcoholic fatty liver disease. Mil Med Res. 2023;10:53.\u003c/span\u003e\u003c/li\u003e\u003cli\u003e\u003cspan\u003eDong J, et al. ACACA reduces lipid accumulation through dual regulation of lipid metabolism and mitochondrial function via AMPK- PPARα- CPT1A axis. J Transl Med. 2024;22:196.\u003c/span\u003e\u003c/li\u003e\u003cli\u003e\u003cspan\u003eDonnelly KL, et al. Sources of fatty acids stored in liver and secreted via lipoproteins in patients with nonalcoholic fatty liver disease. J Clin Invest. 2005;115:1343\u0026ndash;51.\u003c/span\u003e\u003c/li\u003e\u003cli\u003e\u003cspan\u003eFalcon A, et al. FATP2 is a hepatic fatty acid transporter and peroxisomal very long-chain acyl-CoA synthetase. Am J Physiol Endocrinol Metab. 2010;299:E384\u0026ndash;93.\u003c/span\u003e\u003c/li\u003e\u003cli\u003e\u003cspan\u003eFDA. 2005. Guidance for Industry: Estimating the Maximum Safe Starting Dose in Initial Clinical Trials for Therapeutics in Adult Healthy Volunteers Vol., F.a.D. Administration, ed.^eds.\u003c/span\u003e\u003c/li\u003e\u003cli\u003e\u003cspan\u003eFerr\u0026eacute; P, Foufelle F. SREBP-1c transcription factor and lipid homeostasis: clinical perspective. Horm Res. 2007;68:72\u0026ndash;82.\u003c/span\u003e\u003c/li\u003e\u003cli\u003e\u003cspan\u003eFitzgerald PB, et al. The relationship of changes in leptin, neuropeptide Y and reproductive hormones to antipsychotic induced weight gain. Hum Psychopharmacol. 2003;18:551\u0026ndash;7.\u003c/span\u003e\u003c/li\u003e\u003cli\u003e\u003cspan\u003eGavrilova O, et al. Liver peroxisome proliferator-activated receptor gamma contributes to hepatic steatosis, triglyceride clearance, and regulation of body fat mass. J Biol Chem. 2003;278:34268\u0026ndash;76.\u003c/span\u003e\u003c/li\u003e\u003cli\u003e\u003cspan\u003eGebhardt S, et al. Antipsychotic-induced body weight gain: predictors and a systematic categorization of the long-term weight course. J Psychiatr Res. 2009;43:620\u0026ndash;6.\u003c/span\u003e\u003c/li\u003e\u003cli\u003e\u003cspan\u003eGriffin MJ, Sul HS. Insulin regulation of fatty acid synthase gene transcription: roles of USF and SREBP-1c. IUBMB Life. 2004;56:595\u0026ndash;600.\u003c/span\u003e\u003c/li\u003e\u003cli\u003e\u003cspan\u003eGuo J, et al. Upstream stimulating factor 1 suppresses autophagy and hepatic lipid droplet catabolism by activating mTOR. FEBS Lett. 2018;592:2725\u0026ndash;38.\u003c/span\u003e\u003c/li\u003e\u003cli\u003e\u003cspan\u003eHavel RJ. Caloric homeostasis and disorders of fuel transport. N Engl J Med. 1972;287:1186\u0026ndash;92.\u003c/span\u003e\u003c/li\u003e\u003cli\u003e\u003cspan\u003eHouten SM, et al. The Biochemistry and Physiology of Mitochondrial Fatty Acid β-Oxidation and Its Genetic Disorders. Annu Rev Physiol. 2016;78:23\u0026ndash;44.\u003c/span\u003e\u003c/li\u003e\u003cli\u003e\u003cspan\u003eJensen-Urstad AP, Semenkovich CF. Fatty acid synthase and liver triglyceride metabolism: housekeeper or messenger? Biochim Biophys Acta. 2012;1821:747\u0026ndash;53.\u003c/span\u003e\u003c/li\u003e\u003cli\u003e\u003cspan\u003eJiang T, et al. Up-regulation of hepatic fatty acid transporters and inhibition/down-regulation of hepatic OCTN2 contribute to olanzapine-induced liver steatosis. Toxicol Lett. 2019;316:183\u0026ndash;93.\u003c/span\u003e\u003c/li\u003e\u003cli\u003e\u003cspan\u003eJoseph SB, et al. Direct and indirect mechanisms for regulation of fatty acid synthase gene expression by liver X receptors. J Biol Chem. 2002;277:11019\u0026ndash;25.\u003c/span\u003e\u003c/li\u003e\u003cli\u003e\u003cspan\u003eKasper P et al. 2021. Maternal Exercise Mediates Hepatic Metabolic Programming via Activation of AMPK-PGC1α Axis in the Offspring of Obese Mothers. Cells. 10.\u003c/span\u003e\u003c/li\u003e\u003cli\u003e\u003cspan\u003eKawanishi N et al. 2018. Exercise training suppresses scavenger receptor CD36 expression in kupffer cells of nonalcoholic steatohepatitis model mice. Physiol Rep 6, e13902.\u003c/span\u003e\u003c/li\u003e\u003cli\u003e\u003cspan\u003eKelly DL, Conley RR, Tamminga CA. Differential olanzapine plasma concentrations by sex in a fixed-dose study. Schizophr Res. 1999;40:101\u0026ndash;4.\u003c/span\u003e\u003c/li\u003e\u003cli\u003e\u003cspan\u003eKersten S. Integrated physiology and systems biology of PPARα. Mol Metab. 2014;3:354\u0026ndash;71.\u003c/span\u003e\u003c/li\u003e\u003cli\u003e\u003cspan\u003eKlau J et al. 2024. Antipsychotic prescribing patterns in children and adolescents attending Australian general practice in 2011 and 2017. JCPP Adv 4, e12208.\u003c/span\u003e\u003c/li\u003e\u003cli\u003e\u003cspan\u003eKurosaka Y et al. 2021. Protective Effects of Voluntary Exercise on Hepatic Fat Accumulation Induced by Dietary Restriction in Zucker Fatty Rats. Int J Mol Sci 22.\u003c/span\u003e\u003c/li\u003e\u003cli\u003e\u003cspan\u003eLee YK, et al. Hepatic lipid homeostasis by peroxisome proliferator-activated receptor gamma 2. Liver Res. 2018;2:209\u0026ndash;15.\u003c/span\u003e\u003c/li\u003e\u003cli\u003e\u003cspan\u003eLian J, et al. Risperidone-induced weight gain and reduced locomotor activity in juvenile female rats: The role of histaminergic and NPY pathways. Pharmacol Res. 2015;95\u0026ndash;96:20\u0026ndash;6.\u003c/span\u003e\u003c/li\u003e\u003cli\u003e\u003cspan\u003eLiu X, et al. Brown adipose tissue activity is modulated in olanzapine-treated young rats by simvastatin. BMC Pharmacol Toxicol. 2020a;21:48.\u003c/span\u003e\u003c/li\u003e\u003cli\u003e\u003cspan\u003eLiu Y, et al. Hepatic Slug epigenetically promotes liver lipogenesis, fatty liver disease, and type 2 diabetes. J Clin Invest. 2020b;130:2992\u0026ndash;3004.\u003c/span\u003e\u003c/li\u003e\u003cli\u003e\u003cspan\u003eMashek DG. Hepatic fatty acid trafficking: multiple forks in the road. Adv Nutr. 2013;4:697\u0026ndash;710.\u003c/span\u003e\u003c/li\u003e\u003cli\u003e\u003cspan\u003eMay FJ, et al. Lipidomic Adaptations in White and Brown Adipose Tissue in Response to Exercise Demonstrate Molecular Species-Specific Remodeling. Cell Rep. 2017;18:1558\u0026ndash;72.\u003c/span\u003e\u003c/li\u003e\u003cli\u003e\u003cspan\u003eMiles JM, et al. Systemic and forearm triglyceride metabolism: fate of lipoprotein lipase-generated glycerol and free fatty acids. Diabetes. 2004;53:521\u0026ndash;7.\u003c/span\u003e\u003c/li\u003e\u003cli\u003e\u003cspan\u003eMiquilena-Colina ME, et al. Hepatic fatty acid translocase CD36 upregulation is associated with insulin resistance, hyperinsulinaemia and increased steatosis in non-alcoholic steatohepatitis and chronic hepatitis C. Gut. 2011;60:1394\u0026ndash;402.\u003c/span\u003e\u003c/li\u003e\u003cli\u003e\u003cspan\u003eMorrato EH, et al. Metabolic Screening in Children Receiving Antipsychotic Drug Treatment. Arch Pediatr Adolesc Med. 2010;164:344\u0026ndash;51.\u003c/span\u003e\u003c/li\u003e\u003cli\u003e\u003cspan\u003eMorris EM, et al. PGC-1α overexpression results in increased hepatic fatty acid oxidation with reduced triacylglycerol accumulation and secretion. Am J Physiol Gastrointest Liver Physiol. 2012;303:G979\u0026ndash;92.\u003c/span\u003e\u003c/li\u003e\u003cli\u003e\u003cspan\u003eOh KJ, et al. Atypical antipsychotic drugs perturb AMPK-dependent regulation of hepatic lipid metabolism. Am J Physiol Endocrinol Metab. 2011;300:E624\u0026ndash;32.\u003c/span\u003e\u003c/li\u003e\u003cli\u003e\u003cspan\u003ePark H, et al. Coordinate regulation of malonyl-CoA decarboxylase, sn-glycerol-3-phosphate acyltransferase, and acetyl-CoA carboxylase by AMP-activated protein kinase in rat tissues in response to exercise. J Biol Chem. 2002;277:32571\u0026ndash;7.\u003c/span\u003e\u003c/li\u003e\u003cli\u003e\u003cspan\u003ePenninx B, Lange SMM. Metabolic syndrome in psychiatric patients: overview, mechanisms, and implications. Dialogues Clin Neurosci. 2018;20:63\u0026ndash;73.\u003c/span\u003e\u003c/li\u003e\u003cli\u003e\u003cspan\u003ePillinger T, et al. Comparative effects of 18 antipsychotics on metabolic function in patients with schizophrenia, predictors of metabolic dysregulation, and association with psychopathology: a systematic review and network meta-analysis. Lancet Psychiatry. 2020;7:64\u0026ndash;77.\u003c/span\u003e\u003c/li\u003e\u003cli\u003e\u003cspan\u003ePozzi M, et al. Olanzapine, risperidone and ziprasidone differently affect lysosomal function and autophagy, reflecting their different metabolic risk in patients. Transl Psychiatry. 2024;14:13.\u003c/span\u003e\u003c/li\u003e\u003cli\u003e\u003cspan\u003eRazani B, et al. Caveolin-1-deficient Mice Are Lean, Resistant to Diet-induced Obesity, and Show Hypertriglyceridemia with Adipocyte Abnormalities*. J Biol Chem. 2002;277:8635\u0026ndash;47.\u003c/span\u003e\u003c/li\u003e\u003cli\u003e\u003cspan\u003eReagan-Shaw S, Nihal M, Ahmad N. Dose translation from animal to human studies revisited. FASEB J. 2008;22:659\u0026ndash;61.\u003c/span\u003e\u003c/li\u003e\u003cli\u003e\u003cspan\u003eReid BN, et al. Hepatic overexpression of hormone-sensitive lipase and adipose triglyceride lipase promotes fatty acid oxidation, stimulates direct release of free fatty acids, and ameliorates steatosis. J Biol Chem. 2008;283:13087\u0026ndash;99.\u003c/span\u003e\u003c/li\u003e\u003cli\u003e\u003cspan\u003eRosa-Caldwell ME, et al. Moderate physical activity promotes basal hepatic autophagy in diet-induced obese mice. Appl Physiol Nutr Metab. 2017;42:148\u0026ndash;56.\u003c/span\u003e\u003c/li\u003e\u003cli\u003e\u003cspan\u003eSu Y, et al. Epigenetic histone modulations of PPARγ and related pathways contribute to olanzapine-induced metabolic disorders. Pharmacol Res. 2020;155:104703.\u003c/span\u003e\u003c/li\u003e\u003cli\u003e\u003cspan\u003eSu Y et al. 2023. Epigenetic Histone Methylation of PPARγ and CPT1A Signaling Contributes to Betahistine Preventing Olanzapine-Induced Dyslipidemia. Int J Mol Sci 24.\u003c/span\u003e\u003c/li\u003e\u003cli\u003e\u003cspan\u003eSylvester E, et al. Exercise intervention for preventing risperidone-induced dyslipidemia and gluco-metabolic disorders in female juvenile rats. Pharmacol Biochem Behav. 2020;199:173064.\u003c/span\u003e\u003c/li\u003e\u003cli\u003e\u003cspan\u003eTakami G, et al. Effects of atypical antipsychotics and haloperidol on PC12 cells: only aripiprazole phosphorylates AMP-activated protein kinase. J Neural Transm (Vienna). 2010;117:1139\u0026ndash;53.\u003c/span\u003e\u003c/li\u003e\u003cli\u003e\u003cspan\u003eTontonoz P, et al. PPARgamma promotes monocyte/macrophage differentiation and uptake of oxidized LDL. Cell. 1998;93:241\u0026ndash;52.\u003c/span\u003e\u003c/li\u003e\u003cli\u003e\u003cspan\u003eVega RB, Huss JM, Kelly DP. The coactivator PGC-1 cooperates with peroxisome proliferator-activated receptor alpha in transcriptional control of nuclear genes encoding mitochondrial fatty acid oxidation enzymes. Mol Cell Biol. 2000;20:1868\u0026ndash;76.\u003c/span\u003e\u003c/li\u003e\u003cli\u003e\u003cspan\u003eWang Y et al. 2020. PPARs as Metabolic Regulators in the Liver: Lessons from Liver-Specific PPAR-Null Mice. Int J Mol Sci 21.\u003c/span\u003e\u003c/li\u003e\u003cli\u003e\u003cspan\u003eWooten JS, et al. The effects of voluntary wheel running during weight-loss on biomarkers of hepatic lipid metabolism and inflammation in C57Bl/6J mice. Curr Res Physiol. 2022;5:63\u0026ndash;72.\u003c/span\u003e\u003c/li\u003e\u003cli\u003e\u003cspan\u003eWu B, et al. Aerobic exercise promotes the expression of ATGL and attenuates inflammation to improve hepatic steatosis via lncRNA SRA. Sci Rep. 2022;12:5370.\u003c/span\u003e\u003c/li\u003e\u003cli\u003e\u003cspan\u003eXu X, et al. Transcriptional activation of Fsp27 by the liver-enriched transcription factor CREBH promotes lipid droplet growth and hepatic steatosis. Hepatology. 2015;61:857\u0026ndash;69.\u003c/span\u003e\u003c/li\u003e\u003cli\u003e\u003cspan\u003eYi W, et al. Kidney plays an important role in ketogenesis induced by risperidone and voluntary exercise in juvenile female rats. Psychiatry Res. 2021;305:114196.\u003c/span\u003e\u003c/li\u003e\u003cli\u003e\u003cspan\u003eZhu Q, et al. Prevalence and clinical correlates of abnormal lipid metabolism in first-episode and drug-na\u0026iuml;ve patients with major depressive disorder with abnormal glucose metabolism. Sci Rep. 2023;13:8078.\u003c/span\u003e\u003c/li\u003e\u003c/ol\u003e"}],"fulltextSource":"","fullText":"","funders":[],"hasAdminPriorityOnWorkflow":false,"hasManuscriptDocX":true,"hasOptedInToPreprint":true,"hasPassedJournalQc":"","hasAnyPriority":false,"hideJournal":true,"highlight":"","institution":"","isAcceptedByJournal":false,"isAuthorSuppliedPdf":false,"isDeskRejected":"","isHiddenFromSearch":false,"isInQc":false,"isInWorkflow":false,"isPdf":false,"isPdfUpToDate":true,"isWithdrawnOrRetracted":false,"journal":{"display":true,"email":"[email protected]","identity":"researchsquare","isNatureJournal":false,"hasQc":true,"allowDirectSubmit":true,"externalIdentity":"","sideBox":"","snPcode":"","submissionUrl":"/submission","title":"Research Square","twitterHandle":"researchsquare","acdcEnabled":true,"dfaEnabled":false,"editorialSystem":"","reportingPortfolio":"","inReviewEnabled":false,"inReviewRevisionsEnabled":true},"keywords":"Antipsychotic drug, Exercise, Metabolic side effect, Lipid metabolism, Juvenile","lastPublishedDoi":"10.21203/rs.3.rs-7309963/v1","lastPublishedDoiUrl":"https://doi.org/10.21203/rs.3.rs-7309963/v1","license":{"name":"CC BY 4.0","url":"https://creativecommons.org/licenses/by/4.0/"},"manuscriptAbstract":"\u003ch2\u003eObjectives\u003c/h2\u003e\u003cp\u003eRisperidone is a commonly used antipsychotic drug in juveniles, but with serious metabolic side-effects. Previous studies found that exercise reduced plasma triglyceride level and adipose accumulation caused by risperidone. This study elucidated the underlying mechanisms.\u003c/p\u003e\u003ch2\u003eMethods\u003c/h2\u003e\u003cp\u003eFemale juvenile rats were randomly allocated into Vehicle\u0026thinsp;+\u0026thinsp;Sedentary, Risperidone (0.9mg/kg; twice per day)\u0026thinsp;+\u0026thinsp;Sedentary, Vehicle\u0026thinsp;+\u0026thinsp;Exercise (3-hour voluntary access to a running wheel/day), and Risperidone\u0026thinsp;+\u0026thinsp;Exercise groups (n\u0026thinsp;=\u0026thinsp;8/group). After 4-week treatment, the liver was harvested for subsequent examination.\u003c/p\u003e\u003ch2\u003eResults\u003c/h2\u003e\u003cp\u003e(1) Lipogenesis: Protein levels of FAS and USF1 were raised in the risperidone-treated sedentary group, which was decreased by exercise. The pAMPK/AMPK ratio was upregulated by exercise. (2) Lipid uptake/storage: Risperidone-induced upregulations of PPARγ and CD36 were downregulated by exercise. FSP27 expression was decreased by exercise. (3) Lipolysis/β-oxidation: Hepatic protein levels of ATGL and HSL in the Risperidone\u0026thinsp;+\u0026thinsp;Exercise group were larger than Risperidone\u0026thinsp;+\u0026thinsp;Sedentary group. Reduced PGC1α expression was found in the risperidone-only group, which was reversed by exercise.\u003c/p\u003e\u003ch2\u003eConclusion\u003c/h2\u003e\u003cp\u003eRisperidone enhanced fatty acid synthesis via the hepatic USF1/FAS signaling pathway and to augment fatty acid uptake through the PPARγ/CD36 pathway, while simultaneously diminishing β-oxidation by down-regulating hepatic PGC1α expression. Conversely, voluntary exercise intervention counteracted these effects, thereby ameliorating the lipid imbalances induced by risperidone.\u003c/p\u003e","manuscriptTitle":"The effect and mechanisms of risperidone and voluntary exercise intervention on hepatic lipid metabolism in juvenile female rats","msid":"","msnumber":"","nonDraftVersions":[{"code":1,"date":"2025-08-21 10:30:09","doi":"10.21203/rs.3.rs-7309963/v1","editorialEvents":[{"type":"communityComments","content":0}],"status":"published","journal":{"display":true,"email":"[email protected]","identity":"researchsquare","isNatureJournal":false,"hasQc":true,"allowDirectSubmit":true,"externalIdentity":"","sideBox":"","snPcode":"","submissionUrl":"/submission","title":"Research Square","twitterHandle":"researchsquare","acdcEnabled":true,"dfaEnabled":false,"editorialSystem":"","reportingPortfolio":"","inReviewEnabled":false,"inReviewRevisionsEnabled":true}}],"origin":"","ownerIdentity":"53061a63-22b3-46e9-be99-c4e6a3c96d36","owner":[],"postedDate":"August 21st, 2025","published":true,"recentEditorialEvents":[],"rejectedJournal":[],"revision":"","amendment":"","status":"posted","subjectAreas":[],"tags":[],"updatedAt":"2025-09-01T17:53:23+00:00","versionOfRecord":[],"versionCreatedAt":"2025-08-21 10:30:09","video":"","vorDoi":"","vorDoiUrl":"","workflowStages":[]},"version":"v1","identity":"rs-7309963","journalConfig":"researchsquare"},"__N_SSP":true},"page":"/article/[identity]/[[...version]]","query":{"redirect":"/article/rs-7309963","identity":"rs-7309963","version":["v1"]},"buildId":"8U1c8b4HqxoKbykW_rLl7","isFallback":false,"isExperimentalCompile":false,"dynamicIds":[84888],"gssp":true,"scriptLoader":[]}