Long-term offspring loss in lactating rats: Neurobiological and emotional consequences in a novel animal model

Long-term offspring loss in lactating rats: Neurobiological and emotional consequences in a novel animal model | bioRxiv /* */ /* */ <!-- <!-- /*! * yepnope1.5.4 * (c) WTFPL, GPLv2 */ (function(a,b,c){function d(a){return"[object Function]"==o.call(a)}function e(a){return"string"==typeof a}function f(){}function g(a){return!a||"loaded"==a||"complete"==a||"uninitialized"==a}function h(){var a=p.shift();q=1,a?a.t?m(function(){("c"==a.t?B.injectCss:B.injectJs)(a.s,0,a.a,a.x,a.e,1)},0):(a(),h()):q=0}function i(a,c,d,e,f,i,j){function k(b){if(!o&&g(l.readyState)&&(u.r=o=1,!q&&h(),l.onload=l.onreadystatechange=null,b)){"img"!=a&&m(function(){t.removeChild(l)},50);for(var d in y[c])y[c].hasOwnProperty(d)&&y[c][d].onload()}}var j=j||B.errorTimeout,l=b.createElement(a),o=0,r=0,u={t:d,s:c,e:f,a:i,x:j};1===y[c]&&(r=1,y[c]=[]),"object"==a?l.data=c:(l.src=c,l.type=a),l.width=l.height="0",l.onerror=l.onload=l.onreadystatechange=function(){k.call(this,r)},p.splice(e,0,u),"img"!=a&&(r||2===y[c]?(t.insertBefore(l,s?null:n),m(k,j)):y[c].push(l))}function j(a,b,c,d,f){return q=0,b=b||"j",e(a)?i("c"==b?v:u,a,b,this.i++,c,d,f):(p.splice(this.i++,0,a),1==p.length&&h()),this}function k(){var a=B;return a.loader={load:j,i:0},a}var l=b.documentElement,m=a.setTimeout,n=b.getElementsByTagName("script")[0],o={}.toString,p=[],q=0,r="MozAppearance"in l.style,s=r&&!!b.createRange().compareNode,t=s?l:n.parentNode,l=a.opera&&"[object Opera]"==o.call(a.opera),l=!!b.attachEvent&&!l,u=r?"object":l?"script":"img",v=l?"script":u,w=Array.isArray||function(a){return"[object Array]"==o.call(a)},x=[],y={},z={timeout:function(a,b){return b.length&&(a.timeout=b[0]),a}},A,B;B=function(a){function b(a){var a=a.split("!"),b=x.length,c=a.pop(),d=a.length,c={url:c,origUrl:c,prefixes:a},e,f,g;for(f=0;f<d;f++)g=a[f].split("="),(e=z[g.shift()])&&(c=e(c,g));for(f=0;f<b;f++)c=x[f](c);return c}function g(a,e,f,g,h){var i=b(a),j=i.autoCallback;i.url.split(".").pop().split("?").shift(),i.bypass||(e&&(e=d(e)?e:e[a]||e[g]||e[a.split("/").pop().split("?")[0]]),i.instead?i.instead(a,e,f,g,h):(y[i.url]?i.noexec=!0:y[i.url]=1,f.load(i.url,i.forceCSS||!i.forceJS&&"css"==i.url.split(".").pop().split("?").shift()?"c":c,i.noexec,i.attrs,i.timeout),(d(e)||d(j))&&f.load(function(){k(),e&&e(i.origUrl,h,g),j&&j(i.origUrl,h,g),y[i.url]=2})))}function h(a,b){function c(a,c){if(a){if(e(a))c||(j=function(){var a=[].slice.call(arguments);k.apply(this,a),l()}),g(a,j,b,0,h);else if(Object(a)===a)for(n in m=function(){var b=0,c;for(c in a)a.hasOwnProperty(c)&&b++;return b}(),a)a.hasOwnProperty(n)&&(!c&&!--m&&(d(j)?j=function(){var a=[].slice.call(arguments);k.apply(this,a),l()}:j[n]=function(a){return function(){var b=[].slice.call(arguments);a&&a.apply(this,b),l()}}(k[n])),g(a[n],j,b,n,h))}else!c&&l()}var h=!!a.test,i=a.load||a.both,j=a.callback||f,k=j,l=a.complete||f,m,n;c(h?a.yep:a.nope,!!i),i&&c(i)}var i,j,l=this.yepnope.loader;if(e(a))g(a,0,l,0);else if(w(a))for(i=0;i (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];var j=d.createElement(s);var dl=l!='dataLayer'?'&l='+l:'';j.src='//www.googletagmanager.com/gtm.js?id='+i+dl;j.type='text/javascript';j.async=true;f.parentNode.insertBefore(j,f);})(window,document,'script','dataLayer','GTM-M677548'); Skip to main content Home About Submit ALERTS / RSS Search for this keyword Advanced Search New Results Long-term offspring loss in lactating rats: Neurobiological and emotional consequences in a novel animal model View ORCID Profile Luisa Demarchi , View ORCID Profile Alice Sanson , Anna-Lena Boos , View ORCID Profile Oliver J. Bosch doi: https://doi.org/10.1101/2025.02.17.634946 Luisa Demarchi 1 Department of Behavioural and Molecular Neurobiology, Regensburg Center for Neuroscience, University of Regensburg , 93053 Regensburg, Germany 2 Department of Biological Sciences, Fordham University , Bronx, NY, USA Find this author on Google Scholar Find this author on PubMed Search for this author on this site ORCID record for Luisa Demarchi Alice Sanson 1 Department of Behavioural and Molecular Neurobiology, Regensburg Center for Neuroscience, University of Regensburg , 93053 Regensburg, Germany Find this author on Google Scholar Find this author on PubMed Search for this author on this site ORCID record for Alice Sanson Anna-Lena Boos 1 Department of Behavioural and Molecular Neurobiology, Regensburg Center for Neuroscience, University of Regensburg , 93053 Regensburg, Germany Find this author on Google Scholar Find this author on PubMed Search for this author on this site Oliver J. Bosch 1 Department of Behavioural and Molecular Neurobiology, Regensburg Center for Neuroscience, University of Regensburg , 93053 Regensburg, Germany Find this author on Google Scholar Find this author on PubMed Search for this author on this site ORCID record for Oliver J. Bosch For correspondence: oliver.bosch{at}ur.de Abstract Full Text Info/History Metrics Preview PDF Abstract The maternal bond is a vital social connection that supports the survival and well-being of both the caregiver and offspring. Disruption of this bond, particularly following offspring loss, can result in profound trauma with long-lasting consequences. While considerable research has focused on the impact of maternal separation on offspring development, the biological effects of offspring loss on the mother remain largely unexplored. In this study, we examined the long-term effects of offspring loss on neuroplasticity, the oxytocin (OXT) and corticotropin-releasing factor (CRF) systems, and stress-coping behaviors in Sprague-Dawley rat mothers. We examined two groups of lactating mothers: (I) a control group, in which dams remained with their pups until natural weaning, and (II) a separated group, in which all offspring were removed on lactation day 1 and the mothers experienced offspring loss until the time corresponding to weaning (19 days). Our results reveal that pup removal increased OXT receptor binding and reduced dendritic spine density in limbic brain regions, without altering estrogen receptor α or calbindin cell expression. Separated mothers additionally showed elevated plasma corticosterone levels and increased passive stress-coping behaviors in the forced swim test. Remarkably, passive stress-coping behavior was rescued by central CRF receptor blockade but not by OXT treatment, indicating that the CRF system plays a central role in the distress response to offspring loss. These findings establish the rat as a novel animal model for maternal distress, provide new insights into the complex neurobiology of grief, and suggests potential directions for future studies. Introduction The maternal-infant bond is among the strongest social bonds in mammals 1 . In humans, disruption of this bond such as through offspring loss, elicits grief-like reactions in bereaved mothers, encompassing a range of emotional, cognitive, and behavioral responses, including depression and altered stress response. These dysregulations can progress to prolonged grief disorder (PGD) 2 – 4 . Despite the clinical relevance, the neurobiological consequences of offspring loss on the maternal brain -particularly in relation to neuroplasticity and the oxytocin (OXT) and corticotropin-releasing factor (CRF) systems-remain poorly understood. Rodent studies have been instrumental in characterizing maternal bonding. Reduced maternal care in rats negatively impacts the offspring development and acts as emotional trauma (for review see 5 ). Importantly, repeated offspring separation has been shown to induce significant behavioral consequences in mothers 6 -, 9 . However, repeated separations do not fully model the experience of complete offspring loss -a scenario more analogous to human maternal grief. Only a limited number of studies have specifically investigated permanent offspring loss in rat mothers 10 , 11 (for review see 12 ). Our previous work demonstrates that offspring loss immediately after birth alters maternal brain function and stress-coping behavior during the first postpartum week, including heightened passive stress-coping, altered neuronal activity, and decreased OXT receptor (OXT-R) binding in the central amygdala (CeA) 13 . Building on these findings, we hypothesized that prolonged offspring loss over 19 days in rats could cause enduring neurobiological changes paralleling aspects of human grief 12 , with particular emphasis on the CRF and OXT systems. Disruption of social bonds in rodents affects neurotransmitter systems, neuroplasticity and behavior 14 , 15 . For example, in the monogamous and biparental prairie vole ( Microtus ochrogaster ) partner loss enhances CRF system signaling, reduces OXT signaling specifically in the nucleus accumbens (NAcc), and alters microglial activity and morphology in a brain region- and sex-specific manner 16 – 18 . The CRF system, a key mediator of stress and depression, and has been extensively studied in both humans and rodents. Dysregulated CRF system activity is strongly linked to mood disorders 19 – 24 . CRF ligands (CRF, Urocortin 1-3) and their receptors (CRF-R1 and CRF-R2) are widely expressed both peripherally and centrally, and regulate physiological, autonomic, and behavioral responses to stress 20 , 25 . Elevated CRF levels in the cerebrospinal fluid and increased CRF-R binding have been reported in depressed patients 26 , consistent with upregulated CRF system activity 27 , 28 . In rats, CRF overexpression induces depressive-like symptoms, altered hypothalamic-pituitary-adrenal (HPA) axis function, and behavioral changes 29 , while chronic stress can lead to CRF system sensitization 30 . OXT, another key neuropeptide in social bonding, is essential for maternal bonding 31 – 33 , and other forms of social affiliation 34 . OXT-R are widely distributed throughout the brain, where OXT signaling modulates stress responses and facilitates social behavior 35 . OXT is thought to have antidepressant-like properties, partly by dampening HPA axis reactivity 36 – 38 , making it a candidate target for grief- and depression-related disorders 34 , 35 including postpartum depression 39 . Neuroplasticity is another important factor in grief and depression. Alterations in dendritic spine density, a key structural marker of plasticity, have been implicated in depression 40 , though the mechanisms remain incompletely understood 41 . Neuroplasticity and emotional processing are also regulated by estrogen receptor ESR1 by influencing dendritic spine formation and OXT-R expression 42 – 44 . ESR1 is expressed at high density in calbindin-immunoreactive (ir+) cells of the ventromedial hypothalamus (VMH) 45 , and calbindin-ir+ cells are reduced in the occipital cortex of post-mortem depressed patients 46 . We hypothesized that early loss of offspring would lead to enduring changes in the maternal stress-response, neuroendocrine systems and neuroplasticity. To test this, we studied lactating female Sprague-Dawley rats that experienced one day of motherhood followed by 19 days of offspring loss. We examined stress-coping behavior, HPA axis reactivity, OXT-R binding, dendritic spine density, and ESR1 and calbindin ir+ cells expression in key limbic and maternal brain regions. This work provides novel insights into the neurobiology of maternal distress and highlights the potential therapeutic relevance of targeting central CRF-R signaling. Materials and Methods Animals Sprague-Dawley rats were obtained from Charles River Laboratories (Sulzfeld, Germany) and kept under standard laboratory conditions (12:12h light/dark cycle, lights on at 7 a.m., room temperature 22 ± 1°C, relative humidity 55 ± 5°C) with access to standard rat chow (ssniff-Spezialdiäten GmbH, Soest, Germany) and water ad libitum . After 7 days of habituation, females were mated with sexually experienced male Sprague-Dawley rats in standard laboratory cages (Eurostandard Type IV, 60 cm x 40 cm x 20 cm) for 10 days. From pregnancy day 18 on, pregnant females were single-housed for undisturbed delivery in observational cages (plexiglass, 38 cm x 22 cm x 35 cm); all rats delivered 8-15 pups within 3-4 days after being single-housed. The studies were conducted in accordance with the ARRIVE guidelines, the European regulations of animal experimentation (European Directive 2010/63/EU) and were approved by the local government of Unterfranken (Bavaria, Germany). According to the 3-Rs principles, all efforts were made to minimize the number of animals and their distress or suffering. Study design We investigated two primary groups of lactating females: (I) control mothers, which remained with their pups until natural weaning, and (II) separated mothers, which had all offspring removed on lactation day 1 and experienced offspring loss for 19 days ( Fig. 1 ). The 19-day separation period was selected because it corresponds to the pre-weaning stage, a critical time when the mother–offspring bond remains strong in rats 47 . Our study followed a stepwise approach. First, we assessed the effects of prolonged offspring loss on OXT-R binding, ESR1 and calbindin-immunoreactive (ir⁺) cells in brain regions implicated in maternal behavior. This analysis provided an initial characterization of neuroendocrine adaptations within the maternal brain and helped identify target regions for subsequent investigation. Next, we evaluated dendritic spine density and behavioral outcomes, incorporating an additional group of virgin females to distinguish experience-dependent plasticity in maternal neural circuits. Download figure Open in new tab Figure 1: Schematic timelines summarizing the maternal separation paradigm. Abbreviations: LD, lactation day. Experimental groups Rats were randomly assigned to one of three experimental groups: (I) control lactating mothers (primiparous) that remained with their pups until weaning (lactation day 20, LD20 group), (II) separated mothers ( LD1+19 group; primiparous) whose litters were removed on lactation day 1 (LD1), and (III) virgin females ( virgin group; nulliparous). To standardize litter size, each litter was adjusted to eight pups of mixed sexes on the day of delivery (LD0). In the LD1+19 group, all pups were removed on LD1 at 10:00 a.m., after which the mothers were left undisturbed in their home cages. To control for the effects of single housing, virgin females were housed individually for the same duration as separated mothers. Experimental design Experiment 1 : a first cohort of rats followed the protocol described above and was sacrificed on lactation day 20 (LD20) without further manipulations ( Fig. 1 ). Brains were collected for: (i) OXT-R binding analysis ( LD20 = 6; LD1+19 = 6), (ii) immunofluorescence analysis ( LD20 = 7; LD1+19 = 5), and (iii) dendritic spine analysis ( LD20 = 4; LD1+19 = 4; virgin = 5). Experiment 2: a second cohort ( LD20 = 15; LD1+19 = 15; virgin = 9) was tested for locomotor and anxiety-like behavior in the light–dark box (LDB) on LD19 and in the open field (OF) on LD20. Immediately following the final behavioral test, rats were sacrificed. Experiment 3 : a third cohort ( LD20 = 13; LD1+19 = 14; virgin = 8) was evaluated for stress-coping behavior in the single-session forced swim test (FST) on LD20. Within 5 minutes of test completion, rats were sacrificed, and trunk blood was collected for plasma corticosterone (CORT) analysis. Experiments 4 and 5 : two additional cohorts were prepared following the same protocol. On LD15, all rats underwent stereotaxic implantation of an intracerebroventricular (icv) guide cannula for acute substance infusion. On LD19, they completed the 15-min pre-test session of the two-session forced swim test (FST). On LD20, they were tested in a 10-min session following infusion of either vehicle (VEH) or a pharmacological treatment. In Experiment 4, the CRF-R1/2 antagonist D-Phe was administered ( LD20 VEH = 10; LD1+19 VEH = 8; LD1+19 D-Phe = 9), whereas in Experiment 5, synthetic OXT was infused ( LD20 VEH = 8; LD1+19 VEH = 8; LD1+19 OXT = 7). In both experiments, all rats were sacrificed immediately after the final behavioral test and brains taken to verify cannula placement. Brain analyses OXT-R autoradiography Rats were anesthetized and underwent cardiac perfusion with ice-cold 1x PBS and were decapitated. Brains were flash-frozen in n-methylbutane, and stored at -20 °C until cutting into coronal sections of 16 μm using a cryostat (CM3050S, Leica Microsystem GmbH). For each brain region of interest (bed nucleus of the stria terminalis (BNST), nucleus accumbens shell (NAcc shell), medial preoptic area (MPOA), accessory olfactory bulbs (AOB), central amygdala (CeA), agranular insular cortex (AIP), ventromedial hypothalamus (VMH), lateral septum (LS), prelimbic cortex (PL)) six slices per rat were collected on SUPERFROST microscope slides and stored at -20 °C until further processing. The OXT-R autoradiography was performed following an established protocol 48 . Briefly, the ornithin-vasotocin analogue [125I]-OVTA [d(CH2)5[Tyr(Me)2,Thr4,Orn8,[125I]Tyr9-NH2]; Perkin Elmer, USA) was used as a tracer. First, the slides were thawed and allowed to dry thoroughly at room temperature. The tissue was shortly fixed via 0.1 % PFA, washed 2 x in Tris (50 mM, pH 7.4), covered with the tracer solution (50 mM tracer, 10 mM MgCl2, 0.1 % BSA) for 60 min, washed 3 x in Tris / MgCl2 buffer for 7 min, each, followed by 30 min spinning in Tris / MgCl2. Finally, slides were dipped into water and air dried before being exposed to Biomax MR films (Kodak, Cedex, France) for 15 days. The films were scanned using the EPSON Perfection V800 Scanner (Epson GmbH, Munich, Germany), and the optical density of each region of interest was analyzed using ImageJ 49 by subtracting the background activity as previously described 48 , 50 . The analyses were performed simultaneously for 6 slices per rat and per region. Immunofluorescence staining Rats were anesthetized and perfused with ice-cold 1x PBS and subsequently with 4 % paraformaldehyde (PFA) dissolved in 1x PBS. Brains were removed, fixed overnight in 4 % PFA, and subsequently incubated in 30 % sucrose in 1x PBS until the brain sank. After fixation and cryoprotection in sucrose, brains were flash-frozen in n-methylbutane, and stored at -20 °C until cutting into coronal sections of 16 μm using a cryostat (CM3050S, Leica Microsystem GmbH, Nussloch, Germany). Six consecutive slices containing the VMH region were collected per rat on SUPERFROST microscope slides and stored at -20 °C until further processing. Slides were washed in 1x PBS, permeabilized with 0.3 % Triton, blocked with 5 % normal goat serum (Vector Laboratories, Newark, USA), and incubated overnight at 4 °C with primary antibodies (Merck Millipore, rabbit anti ER alpha 1:2000; Merck, mouse anti-NeuN 1:1000; Synaptic Systems, chicken anti-calbindin 1:2000). The next day, slides went through 1 h incubation with secondary antibodies (Alexa Fluor 488-conjugated anti-rabbit IgG 1:800, Alexa Fluor 594-conjugated anti-mouse IgG 1:800 and Alexa Fluor 649-conjugated anti-chicken IgG 1:800) at room temperature. All slides were finally mounted using ROTI®Mount FluorCare DAPI (Carl Roth, Karlsruhe, Germany). Golgi staining for spine density visualization The spine density was assessed in virgin , LD20 and LD1+19 groups to investiganeuronal plasticity. Firstly, rats were sacrificed with CO 2 and brains were collected. The Rapid GolgiStainTM Kit (FD NeuroTechnologies, Columbia, USA) was used and applied according to the manufacturer’s protocol. Brains were cut into 100 μm slices using a cryostat (CM3050S, Leica Microsystem GmbH) at -28° and placed on gelatin-coated microscope slides (FD NeuroTechnologies, Columbia, USA). After complete drying, Golgi staining was performed, and sections were stored at room temperature in the dark before acquiring images using a brightfield microscope (Leica Thunder DM6 B, Camera Leica DFC9000 GT). ImageJ 49 was used to count the spines of secondary dendrites in the PL, AIP, BLA and VMH. Secondary dendrites from different neurons (between 4 and 12, depending on the brain region) were chosen from 10X overview preview images followed by acquiring 40X focus images for analysis. The ROI of each dendrite was selected, brightness and sharpening function were adjusted. Spines were counted on a 25-30 μm 2 section of the dendrite (each dendrite belonging to a different neuron) and the absolute spine density was calculated by using the following formula 51 : (n = number of spines; μm = dendritic length): . Behavioral experiments Light dark box (LDB) On LD19, between 9:00 a.m. and 12:00 p.m., rats from experiment 2 were tested in the LDB to assess anxiety-like behavior 52 . The apparatus consisted of a light compartment (40 × 50 cm, 180 lux) and a dark compartment (40 × 30 cm, 0 lux) connected by a 7.5 × 7.5 cm opening. At the start of the test, rats were placed in the center of the light box, and behavior was recorded for 10 min using EthoVision XT (Noldus, Wageningen, the Netherlands) by an experimenter blind to group assignment. The following parameters were analyzed: (a) percentage of time spent in the light box, (b) latency to re-enter the light compartment, and (c) locomotor activity. The arena was cleaned with tap water and dried thoroughly between trials. Open-field (OF) On LD20, between 9:00 a.m. and 12:00 p.m., rats from experiment 2 were tested in the OF to evaluate locomotor activity and anxiety-like behavior 53 . The apparatus was an empty rectangular arena (80 × 80 cm). Rats were placed in one corner and allowed to explore freely for 10 min. Behavior was recorded using EthoVision XT, with analysis performed by an experimenter blind to treatment. The following measures were scored: (a) locomotion, (b) velocity, and (c) number of center entries. The arena was cleaned with tap water and dried between trials. Forced swim test (FST) Two versions of the FST were employed to assess passive stress-coping behavior. Rationale: we used a single-session protocol to capture baseline group differences without pre-exposure, whereas the two-session protocol provided higher sensitivity to drug effects -particularly relevant because animals in experiment 4-5 underwent surgery (see below). Single-session FST On LD20, between 9:00 a.m. and 12:00 p.m., rats from experiment 3 were tested in a single 10-min FST session. Each rat was placed in a cylindrical tank (50 cm high, 30 cm diameter) filled with tap water (23 ± 1 °C) at a depth preventing contact with the bottom. Behavior was recorded and analyzed using JWatcher ( https://www.jwatcher.ucla.edu ) by an experimenter blind to group assignment. The total time spent immobile (floating) was quantified as an index of passive stress-coping, which is reminiscent of depressive-like behavior 54 . Two-session FST In experiments 4 and 5, rats were tested using the two-day Porsolt FST, the standard paradigm for evaluating antidepressant treatments 54 . These animals had previously undergone stereotaxic surgery (see details below) and were tested 5 days postoperatively. On LD19, rats underwent a 15-min pre-test session. On LD20, they were exposed to a 10-min test session following intracerebroventricular infusion of vehicle or a pharmacological agent. The two-session design was chosen to stabilize baseline immobility at the pre-test and to maximize the dynamic range for detecting drug-induced reductions at the test while controlling for post-surgical variability. All other experimental conditions and behavioral analyses were identical to those described for the single-session FST. Stereotaxic surgery On LD15, rats of experiments 4 and 5 were implanted with a stainless-steel guide cannula (21-G, length 12 mm), above the right ventricle (1.0 mm posterior, 1.6 mm lateral to bregma, 1.8 mm ventral) 55 . Surgeries were performed under isoflurane anesthesia, following previously described procedures 56 . The guide cannula was closed using a 25-G stainless steel stylet to prevent occlusion. To minimize post-surgical discomfort, all rats received a subcutaneous injection of buprenorphine (0.05 mg/kg, Bayer Vital GmbH, Leverkusen, Germany) prior to surgery. Following surgery, rats were monitored continuously until recovery from anesthesia and then returned to their home cages. Rats were observed daily for general health, body weight, and signs of discomfort or infection throughout the recovery period. Drug administration On LD20, rats of experiment 4 and 5 received a single acute icv infusion 10 min prior to the second session of the two-day forced swim test. Depending on the treatment group, animals were infused with vehicle (VEH; 5 µL sterile Ringer’s solution, pH adjusted to 7.4; Braun, Melsungen, Germany), the human/rat CRF-R1/2 antagonist D-Phe [(D-Phe¹², Nle²¹–³⁸, α-Me-Leu³⁷)-CRF (12–41); 10 µg/5 µL; Bachem, Bubendorf, Switzerland], or synthetic OXT (1 µg/5 µL; Tocris, Nordenstadt, Germany). All doses were selected based on previous studies 56 . Infusions were performed using a 25 G stainless-steel infusion cannula (14 mm length) connected to a 10 µL Hamilton micro syringe via PE-50 tubing (50 cm). The infusion cannula was inserted into the pre-implanted guide cannula and held in place with a piece of silicon tubing. Substances were delivered over 30s, after which the cannula was left in position for an additional 30s to minimize reflux before removal, as previously described 38 . Verification of cannula placement Rats of experiment 4 and 5 were sacrificed immediately following the last behavioral test with CO 2 and blue dye was injected via the infusion system into the guide cannula. Verification of the infusion site of the icv cannula was observed by dye spread throughout the ventricular system. Only animals with correct verified infusion sites were included in the statistical analyses. ELISA for plasma CORT In experiment 3, rats were sacrificed within 5 min after the single-session FST, and approximately 1 mL of trunk blood was collected in EDTA-coated tubes on ice (Sarstedt, Numbrecht, Germany). Samples were centrifuged at 4 °C (10,000 rpm/12298 rcf) for 10 min. Plasma was aliquoted and stored at −20 °C until analysis. CORT concentrations were quantified using a commercially available ELISA kit (Tecan IBL International GmbH, Hamburg, Germany) following the manufacturer’s instructions. All samples were measured in duplicate, and mean values were used for statistical analysis. Statistical analysis All statistical analyses were performed with GraphPad PRISM 9 (GraphPad Software, San Diego, USA). Data were first tested for normality (Shapiro-Wilk or Kolmogorov– Smirnov test) and homogeneity of variance (Brown-Forsythe test). Outliers were identified using the ROUT method. Group comparisons of OXT-R binding, ESR1 and calbindin-ir+ cell counts were performed using two-sample Student’s t -test. Dendritic spine density, plasma CORT concentration, anxiety-like behavior, locomotor activity and stress-coping behavior were analyzed by one-way ANOVA followed by Sidak’s post hoc multiple comparisons. Data are expressed as mean ± SEM; and as significance was set at p ≤ 0.05. Results Increased OXT-R binding in the VMH and PL after offspring loss To determine whether offspring loss alters OXT-R binding within maternal and limbic brain regions, we performed receptor autoradiography in LD20 and LD1+19 mothers. LD1+19 dams exhibited significantly higher OXT-R binding in the ventromedial hypothalamus (VMH; M = 0.257, SD = 0.077) compared to LD20 dams ( M = 0.147, SD = 0.083; t (10) = 2.366, p = 0.039; Fig. 2A ). A similar finding was observed in the prelimbic cortex (PL), where OXT-R binding was slightly elevated in LD1+19 ( M = 0.070, SD = 0.005) relative to LD20 dams ( M = 0.066, SD = 0.002, t (10) = 2.203, p = 0.052; Fig. 2B ). Download figure Open in new tab Figure 2: OXT-R binding in brain regions of the limbic system and maternal network. ( A,B ) Representative coronal brain sections of the VMH and PL showing differences in OXT-R binding. Optical density (arbitrary units) for OXT-R binding in the ( A ) VMH and ( B ) PL. ( C ) Overview heatmap of gray density (arbitrary units) for OXT-R binding in the analyzed brain areas (* p ≤ .05 versus LD20 ). Abbreviations: AIP, agranular insular cortex; AOB, accessory olfactory nuclei; BNST, bed nucleus of the stria terminalis; CeA, central amygdala; LS, lateral septum; MPOA, medial preoptic area; NAcc, nucleus accumbens; PL, prelimbic cortex; VMH, ventral medial hypothalamus. Student’s t Test. Data are expressed as mean ±SEM. * p ≤ .05 versus LD20 . No significant group differences were detected in the other regions analyzed ( Fig. 2C ): MPOA ( t (10) = 0.215, p = 0 .834); BNST ( t (10) = 0.828, p = .427); CeA ( t (10) = 0.061, p = 0.952); OB ( t (10) = 0.595, p < 0 .565); NAcc shell ( t (10) = 1.440, p = 0 .180); AIP ( t (10) = 1.212, p = 0 .254); LS ( t (10) = 0.613, p = 0 .554)]. ESR1 and calbindin ir+ cells in the VMH and PL did not differ between groups To investigate whether the increased OXT-R binding in the VMH and PL (see above) was associated with changes in ESR1 expression, we quantified ESR1- and calbindin-immunoreactive (ir+) cells in the VMH and PL of LD20 and LD1+19 mothers. In the VMH, no significant group differences were detected in the proportion of ESR1-ir+ cells ( t (10) = 0.456, p = 0.658; Fig. 3A, B ) or of calbindin ir+ cells ( t (10) = 0.706, p = 0.496; Fig. 3A, C ). ESR1-ir+ cells were not detectable in the PL (data not shown). Download figure Open in new tab Figure 3: ESR1 and calbindin ir+ cells in the VMH. ( A ) Representative fluorescent images showing DAPI (blue), ESR1 cells (red), calbindin (green) and merged channels in the VMH. Percentage of ( B ) ESR1 and ( C ) calbindin ir+ cells in the VMH. One-way ANOVA. Data are expressed as mean ± SEM. Reduced secondary dendritic spine density in the VMH after offspring loss To assess the impact of offspring loss on neuroplasticity, we quantified secondary dendritic spine density in the VMH, PL, BLA and AIP of virgin , LD20 and LD1+19 females ( Fig. 4A ), based on regions identified in our previous study 13 . Statistical analysis revealed a significant group effect in the VMH (one-way ANOVA: F (2, 10) = 8.734; p = 0.006; Fig. 4B ). Post hoc analyses showed that LD1+19 dams exhibited significantly lower spine density compared to virgin ( p = 0.024) and LD20 mothers ( p = 0.009). In contrast, no significant differences were observed in the other regions examined. No differences in spine density were found for the PL ( virgin : 0.64 ± 0.07; LD20 : 0.53 ± 0.04; LD1+19 : 0.62 ± 0.04; one-way ANOVA: F (2,15) = 0.590, p = 0.356), the BLA ( virgin : 0.61 ± 0.03; LD20 : 0.64 ± 0.009; LD1+19 : 0.61 ± 0.02; one-way ANOVA: F (2,16) = 0.158, p = 0.855), and the AIP ( virgin : 0.62 ± 0.05; LD20 : 0.69 ± 0.04; LD1+19 : 0.68 ± 0.04; one-way ANOVA: F (2,15) = 0.599, p = 0.562). Download figure Open in new tab Figure 4: Secondary dendritic spine density in the VMH. ( A ) Representative coronal whole brain section with Golgi staining and schematic illustration of the neuronal dendritic arbor (Scale bar 10 µm). ( B ) Absolute spine density / mm 2 across experimental groups. ( C ) One-way ANOVA. Data are expressed as mean ± SEM. * p < .05 versus all other groups. Anxiety-like behavior was not affected by offspring loss To examine whether offspring loss altered anxiety-like behavior, virgin , LD20 , and LD1+19 rats were tested in the LDB on LD19. Groups differed in latency to re-enter the light compartment (Kruskal–Wallis test, p = 0.045), with LD20 dams tending to be faster (48.6 s ± 15.9 s) than virgin females (213.5 s ± 77.5 s; Dunn’s test, p = 0.058). LD1+19 dams did not differ from either group (121.9 s ± 31.9 s). No group differences were observed in the time spent in the light compartment ( virgin : 34.0 s ± 6.3 s; LD20 : 36.6 s ± 3.6 s; LD1+19 : 42.0 s ± 5.1 s; one-way ANOVA: F (2,36) = 0.676, p = 0.515) or in locomotor activity within the LDB ( virgin : 3778 cm ± 326 cm; LD20 : 3733 cm ± 194 cm; LD1+19 : 3851 cm ± 312 cm; one-way ANOVA: F (2,36) = 0.054, p = 0.948). Thus, LD1+19 mothers did not differ from either virgin or LD20 mothers in any measure of anxiety-like behavior. Locomotor activity was not affected by offspring loss The same cohort of rats was tested in the OF on LD20. Groups differed significantly in distance travelled (one-way ANOVA: F (2,36) = 3.773, p = 0 .032; Fig. 5A ) and in velocity (one-way ANOVA; [ F (2,36) = 3.788], p = 0.032; Fig. 5B ). Post hoc comparisons revealed that virgin females showed greater locomotion ( p = 0.029) and velocity ( p = 0.028) compared with LD20 dams. LD1+19 mothers did not differ from either group. No significant differences were found in the number of center entries ( virgin : 21.4 ± 2.7; LD20 : 16.3 ± 1.5; LD1+19 : 20.5 ± 2.0; one-way ANOVA: F (2,36) = 1.858, p = 0.170). Overall, offspring loss did not alter locomotor activity. Download figure Open in new tab Figure 5: ( A) Locomotor behavior and (B ) velocity in the OF, ( C ) passive stress-coping behavior in the classic FST, and ( D ) corresponding plasma CORT concentration. ( A ) Total distance traveled in cm and ( B ) mean velocity over the 10-min OF test. ( C ) Percentage of floating during the 10-min classic FST. ( D ) Plasma CORT concentration 15 min after start of the FST. One-way ANOVA. Data are expressed as mean ± SEM. # p < .05 versus LD20 VEH ; * p < .05 versus all other groups. Offspring loss increased passive stress-coping behavior and induced a virgin-like stress response A different cohort of virgin , LD20 and LD1+19 rats was tested for passive stress-coping behavior in the single-session FST on LD20. Groups differed significantly in time spent floating (one-way A N O VA : F (2,32) = 6.233, p = 0.005; Fig. 5C ). LD1+19 dams spent more time floating compared with both virgin ( p = 0.019) and LD20 ( p = 0.014) rats. Stress-induced plasma CORT levels, which were collected 15 min after the start of the FST, also differed across groups ( F (2,28) = 13.50, p = 0.0001; Fig. 5D ). Post hoc comparisons revealed significantly lower CORT concentrations in LD20 compared with virgin ( p = 0.002) and LD1+19 ( p = 0.0001) rats. LD1+19 mothers did not differ from virgin females ( p > 0.05), suggesting that offspring loss abolished the lactation-induced blunting of HPA axis activity. Central CRF-R1/2 blockade normalized passive-stress coping behavior in LD1+19 mothers Following a pre-test on LD19, LD20 and LD1+19 rats were tested in the FST on LD20 after ICV infusion of VEH or the CRF-R1/2 antagonist D-Phe. Groups differed significantly in floating behavior (one-way ANOVA: F (2,23) = 5.828, p = 0.013; Fig. 6A ). LD1+19 VEH mothers spent more time floating than LD20 VEH rats ( p = 0.047), replicating the findings from the single-session FST ( Fig. 5C ). Importantly, floating time was significantly lower in LD1+19 D-Phe rats compared to LD1+19 VEH ( p = 0.014), reaching levels indistinguishable from LD20 VEH ( p > 0.05). Download figure Open in new tab Figure 6: ( A ) Passive stress-coping behavior in the FST following central pharmacological manipulation of the brain CRF and ( B ) OXT systems. ( A ) Percentage of floating during the 10-min modified FST following icv infusion of VEH or the CRF-R1/2 antagonist D-Phe, and ( B ) of VEH or synthetic OXT. One-way ANOVA. Data are expressed as mean ± SEM. * p < .05 versus all other groups; # p < .05 versus LD20 VEH . Central OXT infusion did not alter passive-stress coping behavior in LD1+19 mothers In a different cohort, LD20 and LD1+19 rats were tested in the FST after infusion of VEH or synthetic OXT. Groups differed in floating behavior (one-way ANOVA: F (2,20) = 5.273, p = 0.014; Fig. 6B ). Consistent with previous experiments, LD1+19 VEH rats floated more than LD20 VEH ( p = 0.014). However, infusion of OXT in LD1+19 mothers was not sufficient to alter floating when compared with LD1+19 VEH ( p = 0.08). Discussion The loss of offspring represents a profound life event with strong effects on physiology and behavior. In humans, such experiences are associated with marked changes in emotional and physical health, including a higher risk for prolonged grief disorders 4 , 57 – 59 . To investigate the underlying mechanisms and to identify potential therapeutic targets, appropriate animal models are required. In this study, we build upon our previous work on consequences of short-term separation on rat mothers 13 . We examined the long-term consequences of offspring loss and demonstrate that extended separation manifests in distinct neurobiological and behavioral adaptations. After 19 days of pup removal, mothers exhibited increased OXT-R binding specifically in the VMH and PL, had reduced dendritic spine density in the VMH, and heightened HPA axis reactivity together with increased passive stress-coping behavior. Importantly, central pharmacological blockade of CRF-R1/2 normalized the impaired behavior, whereas OXT infusion did not, highlighting the prominent role of CRF system activity in mediating maternal distress responses due to pup loss. Increased OXT-R binding in the VMH and PL after offspring loss During the peripartum period, the maternal brain undergoes dynamic adaptations in OXT signaling that support the initiation and maintenance of maternal behaviors 60 , 61 . In our study, OXT-R binding was significantly elevated in the VMH and PL following offspring loss, contrary to our initial hypothesis that separation would reduce OXT signaling. The VMH, which contains a high density of OXT-Rs and plays a role in maternal care, aggression, and defensive behaviors 62 – 66 , has not been studied in the context of pup removal. The PL, a region critical for cognitive and emotional regulation 67 , 68 , has also been implicated in maternal care, as OXT-R blockade in this area disrupts caregiving behavior 69 . Thus, the upregulation of OXT-Rs in both the VMH and PL may reflect adaptive—yet potentially insufficient—neurobiological adjustments triggered by permanent offspring removal. Previous studies investigating repeated maternal separation also reported changes in OXT-R binding in the PL and MPOA 6 , 70 , 71 , consistent with the sensitivity of OXT signaling to maternal experience. Reduced maternal interactions, including fewer opportunities for the milk ejection reflex, likely decrease endogenous OXT release 72 , which has been associated with reduced activity of OXT-positive neurons in the PVN 73 . Based on this evidence, we propose that permanent offspring loss alters OXT dynamics, with potentially less OXT release leading to the detected OXT-R upregulation thereby serving as a compensatory adaptation. Future studies should examine brain region-specific OXT release and transcriptional regulation to provide a more complete picture of system-wide neuroendocrine adaptations. ESR1 and calbindin do not mediate OXT-R changes Estrogen has been shown to regulate OXT-R expression via ESR1 and to interact with calbindin 44 , 74 – 76 . To assess whether the OXT-R upregulation observed in our study could be explained by alterations of these factors, we quantified ESR1- and calbindin-immunoreactive (ir⁺) cells in the VMH and PL. No differences were detected in the proportion of ESR1- or calbindin-ir⁺ cells in the VMH between groups, and ESR1 was not detectable in the PL. These results indicate that the increase in OXT-R binding following offspring loss is unlikely to be mediated by changes in ESR1 or calbindin expression. Instead, other neuroendocrine pathways may contribute to the observed receptor alterations. In particular, CRF signaling represents a likely candidate, given its known interactions with the OXT system and its role in stress-related neuroadaptations 77 . Reduced spine density in the VMH after offspring loss Neuroplasticity is a central component of maternal adaptations, with dendritic spine remodeling occurring across pregnancy and the postpartum period 78 . In our study, we observed a selective reduction in secondary dendritic spine density within the VMH after offspring loss, whereas no significant changes were detected in other regions examined. Alterations in dendritic spine density have been widely reported in response to stress and depression in both humans and rodents 79 – 89 . Given the VMH’s established role in the regulation of stress reactivity, social interactions, and aggression 90 – 92 , the observed decrease in spine density may contribute to the stress-response dysregulation observed in separated mothers. Importantly, these structural changes were accompanied by increased OXT-R binding in the VMH, indicating that both cellular and receptor-level adaptations converge in this region. A plausible interpretation is that reduced spine density may reflect altered excitatory connectivity within VMH circuits, while the concurrent increase in OXT-R binding could shift the balance of neuromodulatory input, thereby reshaping local network responsiveness. Rather than acting as a simple compensatory mechanism, these parallel adaptations may represent a reorganization of VMH function following offspring loss, potentially influencing how this region integrates social and stress-related signals. A limitation of the present work is that estrous cycle phases of virgin females were not monitored, which could have influenced spine density measurements. Future studies should therefore control for reproductive state and directly test the functional link between OXT-R signaling and structural remodeling in the VMH to clarify how these adaptations interact to shape maternal behavioral responses. Anxiety-like and locomotor behaviors were unchanged Lactation is often associated with reduced anxiety-like behavior, largely mediated by elevated central OXT signaling 93 , 94 . Consistent with this direction, LD20 dams showed a trend toward faster re-entry versus virgin females, but this did not generalize to other LDB parameters. Crucially, separated mothers ( LD1+19 ) did not differ from LD20 or virgin females on any measure, indicating that offspring loss did not produce robust changes in anxiety-related behavior. The absence of locomotor differences across groups further supports that the FST effects were not secondary to altered mobility or general activity. While the lack of pup-derived sensory stimulation in separated mothers could diminish central OXT release -and thereby remove a potential anxiolytic mechanism 95 -the present data indicates no measurable anxiety-like phenotype in the LDB. Subtle behavioral differences observed in virgin females may reflect their period of social isolation, a known driver of heightened anxiety/hyperactivity in rodents. 96 Offspring loss increased passive stress-coping and HPA axis reactivity Our results show that permanent offspring loss exerted a sustained impact on maternal emotional regulation. In the FST, separated mothers consistently exhibited elevated passive stress-coping, as indicated by increased floating time, compared with both virgin and LD20 dams. This outcome replicates and extends previous findings that prolonged maternal separation enhances passive stress-coping in rat dams¹⁰, ¹¹,¹³. The consistent emergence of this phenotype across independent cohorts emphasizes its robustness and supports the validity of this model for probing maternal adaptations to offspring loss. Importantly, locomotor activity did not differ between LD1+19 and LD20 dams, confirming that the increase in floating reflects a shift in stress-coping strategy rather than reduced motor capacity or energy availability. In addition, physiological analyses aligned with these behavioral findings. LD20 mothers displayed the known dampened HPA axis response, consistent with reduced stress reactivity during the postpartum period 97 . By contrast, LD1+19 mothers exhibited CORT levels similar to virgin females, indicating that offspring loss disrupts the typical stress-buffering adaptations of lactation. This reversion to a virgin-like endocrine profile suggests both heightened vulnerability to acute stressors and the absence of maternal neuroendocrine mechanisms that normally confer resilience during caregiving. Whether this shift reflects the acute impact of pup loss on HPA axis regulation or the lack of sustained maternal experience remains an open question. Together, these findings demonstrate that offspring loss induces a dual effect: an increase in passive stress-coping behavior and the reactivation of heightened HPA axis activity. This behavioral–endocrine combination provides a mechanistic framework for understanding how the maternal brain adapts—or fails to adapt—after loss and reinforces the translational relevance of this model. Central CRF-R1/2 antagonism, but not OXT-R agonism, reversed passive stress-coping During lactation, activity of the brain CRF system is normally dampened, providing resilience against stress and allowing mothers to maintain adaptive caregiving responses 98 . However, chronic or severe stress around parturition can disrupt this regulation, leading to heightened CRF signaling and increased vulnerability to stress 99 , 100 . Furthermore, the CRF system has been implicated in behavioral adaptations following social bond disruption, as shown in prairie voles where partner separation activates CRF-dependent stress responses 101 . In line with this, our results demonstrate that icv administration of the CRF-R1/2 antagonist D-Phe normalized the increased passive stress-coping observed in separated mothers during the FST. These findings indicate that offspring loss induces hyperactivity of the CRF system and that CRF receptor blockade can effectively restore adaptive stress-coping. This interpretation aligns with prior work showing that D-Phe alleviates separation-induced behavioral alterations in both lactating 101 and male prairie voles¹⁶. Given the known functional interactions between CRF and OXT systems 18 , 22 , we also investigated whether enhancing OXT signaling could counteract the behavioral phenotype. Previous studies in prairie voles demonstrated that local OXT infusion into the NAcc shell rescues separation-induced behavioral deficits¹⁸, and work in lactating rats highlights OXT’s regulatory role in stress responses. However, in our study central OXT infusion did not modify passive stress-coping in separated mothers. This lack of effect does not exclude the OXT system as a relevant modulatory pathway; rather, it suggests that global central infusion may not capture the region-specific actions of OXT. Indeed, targeted manipulations in regions such as the PVN 102 or NAcc have produced stronger behavioral effects in other models. Together, our data highlight CRF hyperactivity as a central mechanism in the stress-related consequences of offspring loss, while suggesting that OXT-based interventions may require more precise spatial targeting to be effective. Conclusions This study establishes a translational rat model to investigate the neurobiological consequences of permanent offspring loss. Separated mothers displayed altered stress-coping strategies, elevated HPA axis activity, and structural as well as receptor-level adaptations in limbic brain regions. Specifically, increased OXT-R binding and reduced dendritic spine density were observed in the VMH, identifying this region as a key locus of maternal adaptation to loss. Importantly, pharmacological blockade of CRF-R1/2 signaling restored normal coping behavior, whereas OXT infusion did not, underscoring the CRF system as a primary driver of these outcomes. These findings provide novel insights into the interplay between neuroendocrine signaling, neuroplasticity, and behavioral regulation in the maternal brain. They also suggest that interventions targeting CRF signaling hold particular promise for mitigating the long-term consequences of disrupted maternal experience. Author contributions L.D. designed the study, performed the experiments, analyzed data, and wrote the first draft of the manuscript. A.S. designed the study, performed the experiments and revised the manuscript. A.B. performed the experiments. O.J.B. designed the study, secured funding, supervised the project, and revised the manuscript. All authors contributed to and have approved the final manuscript. Competing interest statement No potential conflict of interest was reported by the authors. Data availability statement The data that support the findings of this study are available from the corresponding author upon reasonable request. Acknowledgements We thank Dr. J. Pawluski and Dr. B. di Benedetto for scientific discussion. We also thank M. Fuchs, A. Havasi, R. Maloumby, M. Schwarz, M. Breitkopf, L. Saller, E. Rocaboy, S. Pena-Pena and T. Gebhardt for excellent technical help. This study was supported by the Deutsche Forschungsgemeinschaft (DFG), through the Neuroscience Graduate Programme “Neurobiology of Emotion Dysfunctions” (GRK 2174, to O.J.B.) and the grant BO1958/8-2 (to O.J.B.). Footnotes Email Luisa Demarchi, ldemarchi{at}fordham.edu , Alice Sanson, alice.sanson{at}ur.de , Anna-Lena Boos, anna-lenaboos{at}gmx.de This version of the manuscript has been revised to update the text and figures. References ↵ Rilling , J. K. & Young , L. J . The biology of mammalian parenting and its effect on offspring social development . Science doi: 10.1126/science.1252723 ( 2014 ). OpenUrl Abstract / FREE Full Text ↵ O’Connor , M. F . Grief: A Brief History of Research on How Body, Mind, and Brain Adapt . Psychosom Med doi: 10.1097/PSY.0000000000000717 ( 2019 ). OpenUrl CrossRef PubMed Alves , R. L. et al. Maternal stress and vulnerability to depression: coping and maternal care strategies and its consequences on adolescent offspring . Transl Psychiatry doi: 10.1038/s41398-022-02220-5 ( 2022 ). OpenUrl CrossRef PubMed ↵ McCarthy , M. C. et al. Prevalence and predictors of parental grief and depression after the death of a child from cancer . J Palliat Med doi: 10.1089/jpm.2010.0037 ( 2010 ). OpenUrl CrossRef PubMed ↵ Nemeroff , C. B . Paradise Lost: The Neurobiological and Clinical Consequences of Child Abuse and Neglect . Neuron doi: 10.1016/j.neuron.2016.01.019 ( 2016 ). OpenUrl CrossRef PubMed ↵ Demarchi , L. , Sanson , A. & Bosch , O. J . Brief versus long maternal separation in lactating rats: Consequences on maternal behavior, emotionality, and brain oxytocin receptor binding . J Neuroendocrinol doi: 10.1111/jne.13252 ( 2023 ). OpenUrl CrossRef Baracz , S. J. , Everett , N. A. , Robinson , K. J. , Campbell , G. R. & Cornish , J. L . Maternal separation changes maternal care, anxiety-like behaviour and expression of paraventricular oxytocin and corticotrophin-releasing factor immunoreactivity in lactating rats . J Neuroendocrinol doi: 10.1111/jne.12861 ( 2020 ). OpenUrl CrossRef PubMed Bolukbas , I. , Mundorf , A. & Freund , N . Maternal separation in rats induces neurobiological and behavioral changes on the maternal side . Sci Rep doi: 10.1038/s41598-020-80087-6 ( 2020 ). OpenUrl CrossRef PubMed ↵ Alves , R. L. , Portugal , C. C. , Summavielle , T. , Barbosa , F. & Magalhaes , A . Maternal separation effects on mother rodents’ behaviour: A systematic review . Neurosci Biobehav Rev doi: 10.1016/j.neubiorev.2019.09.008 ( 2020 ). OpenUrl CrossRef ↵ Pawluski , J. L. , Lieblich , S. E. & Galea , L. A . Offspring-exposure reduces depressive-like behaviour in the parturient female rat . Behav Brain Res doi: 10.1016/j.bbr.2008.08.001 ( 2009 ). OpenUrl CrossRef PubMed ↵ Rincon-Cortes , M. & Grace , A. A . Early Pup Removal Leads to Social Dysfunction and Dopamine Deficit in Late Postpartum Rats: Prevention by Social Support . Front Glob Womens Health doi: 10.3389/fgwh.2021.694808 ( 2021 ). OpenUrl CrossRef ↵ Demarchi , L. , Pawluski , J. L. & Bosch , O. J . The brain oxytocin and corticotropin-releasing factor systems in grieving mothers: What we know and what we need to learn . Peptides doi: 10.1016/j.peptides.2021.170593 ( 2021 ). OpenUrl CrossRef PubMed ↵ Demarchi , L. S. , S; Bosch , O.J. Neurobiological traces of grief: Examining the impact of offspring loss after birth on rat mothers’ brain and stress-coping behavior in the first week postpartum . Neuroscience Applied doi: 10.1016/j.nsa.2024.104065 ( 2024 ). OpenUrl CrossRef PubMed ↵ Shirenova , S. D. , Khlebnikova , N. N. , Narkevich , V. B. , Kudrin , V. S. & Krupina , N. A . Nine-month-long Social Isolation Changes the Levels of Monoamines in the Brain Structures of Rats: A Comparative Study of Neurochemistry and Behavior . Neurochem Res doi: 10.1007/s11064-023-03858-3 ( 2023 ). OpenUrl CrossRef PubMed ↵ Dimonte , S. et al. Social isolation from early life induces anxiety-like behaviors in adult rats: Relation to neuroendocrine and neurochemical dysfunctions . Biomed Pharmacother doi: 10.1016/j.biopha.2022.114181 ( 2023 ). OpenUrl CrossRef PubMed ↵ Bosch , O. J. , Nair , H. P. , Ahern , T. H. , Neumann , I. D. & Young , L. J . The CRF system mediates increased passive stress-coping behavior following the loss of a bonded partner in a monogamous rodent . Neuropsychopharmacology doi: 10.1038/npp.2008.154 ( 2009 ). OpenUrl CrossRef PubMed Web of Science Pohl , T. T. , Jung , O. , Di Benedetto , B. , Young , L. J. & Bosch , O. J . Microglia react to partner loss in a sex- and brain site-specific manner in prairie voles . Brain Behav Immun doi: 10.1016/j.bbi.2021.05.026 ( 2021 ). OpenUrl CrossRef ↵ Bosch , O. J. et al. Oxytocin in the nucleus accumbens shell reverses CRFR2-evoked passive stress-coping after partner loss in monogamous male prairie voles . Psychoneuroendocrinology doi: 10.1016/j.psyneuen.2015.11.011 ( 2016 ). OpenUrl CrossRef PubMed ↵ Fernandez Macedo , G. V. , Cladouchos , M. L. , Sifonios , L. , Cassanelli , P. M. & Wikinski , S. Effects of fluoxetine on CRF and CRF1 expression in rats exposed to the learned helplessness paradigm . Psychopharmacology (Berl ) doi: 10.1007/s00213-012-2859-x ( 2013 ). OpenUrl CrossRef ↵ Binder , E. B. & Nemeroff , C. B . The CRF system, stress, depression and anxiety-insights from human genetic studies . Mol Psychiatry doi: 10.1038/mp.2009.141 ( 2010 ). OpenUrl CrossRef PubMed Web of Science Klampfl , S. M. , Brunton , P. J. , Bayerl , D. S. & Bosch , O. J . CRF-R1 activation in the anterior-dorsal BNST induces maternal neglect in lactating rats via an HPA axis-independent central mechanism . Psychoneuroendocrinology doi: 10.1016/j.psyneuen.2015.11.015 ( 2016 ). OpenUrl CrossRef PubMed ↵ Klampfl , S. M. et al. Maternal stress and the MPOA: Activation of CRF receptor 1 impairs maternal behavior and triggers local oxytocin release in lactating rats . Neuropharmacology doi: 10.1016/j.neuropharm.2018.02.019 ( 2018 ). OpenUrl CrossRef PubMed Sanson , A. , Demarchi , L. , Rocaboy , E. & Bosch , O. J . Increased CRF-R1 transmission in the nucleus accumbens shell facilitates maternal neglect in lactating rats and mediates anxiety-like behaviour in a sex-specific manner . Neuropharmacology doi: 10.1016/j.neuropharm.2024.110256 ( 2025 ). OpenUrl CrossRef PubMed ↵ Carpenter , L. L. et al. Cerebrospinal fluid corticotropin-releasing factor and perceived early-life stress in depressed patients and healthy control subjects . Neuropsychopharmacology doi: 10.1038/sj.npp.1300375 ( 2004 ). OpenUrl CrossRef PubMed Web of Science ↵ Deussing , J. M. & Chen , A . The Corticotropin-Releasing Factor Family: Physiology of the Stress Response . Physiol Rev doi: 10.1152/physrev.00042.2017 ( 2018 ). OpenUrl CrossRef PubMed ↵ Pandey , G. N. , Rizavi , H. S. , Bhaumik , R. & Ren , X . Increased protein and mRNA expression of corticotropin-releasing factor (CRF), decreased CRF receptors and CRF binding protein in specific postmortem brain areas of teenage suicide subjects . Psychoneuroendocrinology doi: 10.1016/j.psyneuen.2019.04.015 ( 2019 ). OpenUrl CrossRef PubMed ↵ Arborelius , L. , Owens , M. J. , Plotsky , P. M. & Nemeroff , C. B . The role of corticotropin-releasing factor in depression and anxiety disorders . J Endocrinol doi: 10.1677/joe.0.1600001 ( 1999 ). OpenUrl Abstract ↵ Davis , E. G. et al. Corticotropin-releasing factor 1 receptor haplotype and cognitive features of major depression . Transl Psychiatry doi: 10.1038/s41398-017-0051-0 ( 2018 ). OpenUrl CrossRef PubMed ↵ Flandreau , E. I. , Ressler , K. J. , Owens , M. J. & Nemeroff , C. B . Chronic overexpression of corticotropin-releasing factor from the central amygdala produces HPA axis hyperactivity and behavioral anxiety associated with gene-expression changes in the hippocampus and paraventricular nucleus of the hypothalamus . Psychoneuroendocrinology doi: 10.1016/j.psyneuen.2011.04.014 ( 2012 ). OpenUrl CrossRef PubMed Web of Science ↵ Herman , J. P. et al. Regulation of the Hypothalamic-Pituitary-Adrenocortical Stress Response . Compr Physiol doi: 10.1002/cphy.c150015 ( 2016 ). OpenUrl CrossRef PubMed ↵ Marlin , B. J. , Mitre , M. , D’Amour J , A. , Chao , M. V. & Froemke , R. C . Oxytocin enables maternal behaviour by balancing cortical inhibition . Nature doi: 10.1038/nature14402 ( 2015 ). OpenUrl CrossRef PubMed Sanson , A. & Bosch , O. J . Dysfunctions of brain oxytocin signaling: Implications for poor mothering . Neuropharmacology doi: 10.1016/j.neuropharm.2022.109049 ( 2022 ). OpenUrl CrossRef PubMed ↵ Carcea , I. et al. Oxytocin neurons enable social transmission of maternal behaviour . Nature doi: 10.1038/s41586-021-03814-7 ( 2021 ). OpenUrl CrossRef PubMed ↵ Menon , R. & Neumann , I. D . Detection, processing and reinforcement of social cues: regulation by the oxytocin system . Nat Rev Neurosci , doi: 10.1038/s41583-023-00759-w ( 2023 ). OpenUrl CrossRef PubMed ↵ Jurek , B. & Neumann , I. D . The Oxytocin Receptor: From Intracellular Signaling to Behavior . Physiol Rev doi: 10.1152/physrev.00031.2017 ( 2018 ). OpenUrl CrossRef PubMed ↵ Arletti , R. & Bertolini , A . Oxytocin acts as an antidepressant in two animal models of depression . Life Sci doi: 10.1016/0024-3205(87)90600-x ( 1987 ). OpenUrl CrossRef Windle , R. J. , Shanks , N. , Lightman , S. L. & Ingram , C. D . Central oxytocin administration reduces stress-induced corticosterone release and anxiety behavior in rats . Endocrinology doi: 10.1210/endo.138.7.5255 ( 1997 ). OpenUrl CrossRef ↵ Neumann , I. D. , Wigger , A. , Torner , L. , Holsboer , F. & Landgraf , R . Brain oxytocin inhibits basal and stress-induced activity of the hypothalamo-pituitary-adrenal axis in male and female rats: partial action within theparaventricular nucleus . J Neuroendocrinol doi: 10.1046/j.1365-2826.2000.00442.x ( 2000 ). OpenUrl CrossRef PubMed Web of Science ↵ Rashidi , M. et al. Peripartum effects of synthetic oxytocin: The good, the bad, and the unknown . Neurosci Biobehav Rev doi: 10.1016/j.neubiorev.2022.104859 ( 2022 ). OpenUrl CrossRef PubMed ↵ Ho , T. C. & King , L. S . Mechanisms of neuroplasticity linking early adversity to depression: developmental considerations . Transl Psychiatry doi: 10.1038/s41398-021-01639-6 ( 2021 ). OpenUrl CrossRef PubMed ↵ Price , R. B. & Duman , R . Neuroplasticity in cognitive and psychological mechanisms of depression: an integrative model . Mol Psychiatry doi: 10.1038/s41380-019-0615-x ( 2020 ). OpenUrl CrossRef PubMed ↵ Sheppard , P. A. S. , Choleris , E. & Galea , L. A. M . Structural plasticity of the hippocampus in response to estrogens in female rodents . Mol Brain doi: 10.1186/s13041-019-0442-7 ( 2019 ). OpenUrl CrossRef PubMed Young , L. J. , Wang , Z. , Donaldson , R. & Rissman , E. F . Estrogen receptor alpha is essential for induction of oxytocin receptor by estrogen . Neuroreport doi: 10.1097/00001756-199803300-00031 ( 1998 ). OpenUrl CrossRef PubMed Web of Science ↵ Jaric , I. , Rocks , D. , Greally , J. M. , Suzuki , M. & Kundakovic , M . Chromatin organization in the female mouse brain fluctuates across the oestrous cycle . Nat Commun doi: 10.1038/s41467-019-10704-0 ( 2019 ). OpenUrl CrossRef PubMed ↵ Mori , H. , Matsuda , K. , Pfaff , D. W. & Kawata , M . A recently identified hypothalamic nucleus expressing estrogen receptor alpha . Proc Natl Acad Sci U S A doi: 10.1073/pnas.0806503105 ( 2008 ). OpenUrl Abstract / FREE Full Text ↵ Maciag , D. et al. Reduced density of calbindin immunoreactive GABAergic neurons in the occipital cortex in major depression: relevance to neuroimaging studies . Biol Psychiatry doi: 10.1016/j.biopsych.2009.10.027 ( 2010 ). OpenUrl CrossRef PubMed Web of Science ↵ Numan , M. & Young , L. J . Neural mechanisms of mother-infant bonding and pair bonding: Similarities, differences, and broader implications . Horm Behav doi: 10.1016/j.yhbeh.2015.05.015 ( 2016 ). OpenUrl CrossRef PubMed ↵ Oliveira , V. E. M. et al. Oxytocin and vasopressin within the ventral and dorsal lateral septum modulate aggression in female rats . Nat Commun doi: 10.1038/s41467-021-23064-5 ( 2021 ). OpenUrl CrossRef PubMed ↵ Schneider , C. A. , Rasband , W. S. & Eliceiri , K.W . NIH Image to ImageJ: 25 years of image analysis . Nat Methods doi: 10.1038/nmeth.2089 ( 2012 ). OpenUrl CrossRef PubMed Web of Science ↵ Bosch , O. J. & Neumann , I. D . Brain vasopressin is an important regulator of maternal behavior independent of dams’ trait anxiety . Proc Natl Acad Sci U S A doi: 10.1073/pnas.0807412105 ( 2008 ). OpenUrl Abstract / FREE Full Text ↵ Gu , L. et al. Long-term in vivo imaging of dendritic spines in the hippocampus reveals structural plasticity . J Neurosci doi: 10.1523/JNEUROSCI.1464-14.2014 ( 2014 ). OpenUrl Abstract / FREE Full Text ↵ Crawley , J. & Goodwin , F. K . Preliminary report of a simple animal behavior model for the anxiolytic effects of benzodiazepines . Pharmacol Biochem Behav doi: 10.1016/0091-3057(80)90067-2 ( 1980 ). OpenUrl CrossRef PubMed Web of Science ↵ Gould T. D. ; Dao D. T. , K. C. E. Mood and Anxiety Related Phenotypes in Mice . Neuromethods doi: 10.1007/978-1-60761-303-9_1 ( 2009 ). OpenUrl CrossRef ↵ Slattery , D. A. & Cryan , J. F . The ups and downs of modelling mood disorders in rodents . ILAR J doi: 10.1093/ilar/ilu026 ( 2014 ). OpenUrl CrossRef PubMed ↵ Paxinos , G. , Watson , C. The Rat Brain in Stereotactic Coordinates . Academic Press , San Diego ( 1998 ). ↵ Klampfl , S. M. , Neumann , I. D. & Bosch , O. J . Reduced brain corticotropin-releasing factor receptor activation is required for adequate maternal care and maternal aggression in lactating rats . Eur J Neurosci doi: 10.1111/ejn.12274 ( 2013 ). OpenUrl CrossRef PubMed ↵ Kersting , A. & Wagner , B . Complicated grief after perinatal loss . Dialogues Clin Neurosci doi: 10.31887/DCNS.2012.14.2/akersting ( 2012 ). OpenUrl CrossRef PubMed Burden , C. et al. From grief, guilt pain and stigma to hope and pride - a systematic review and meta-analysis of mixed-method research of the psychosocial impact of stillbirth . BMC Pregnancy Childbirth doi: 10.1186/s12884-016-0800-8 ( 2016 ). OpenUrl CrossRef PubMed ↵ Huh , H. J. , Huh , S. , Lee , S. H. & Chae , J. H . Unresolved Bereavement and Other Mental Health Problems in Parents of the Sewol Ferry Accident after 18 Months . Psychiatry Investig doi: 10.4306/pi.2017.14.3.231 ( 2017 ). OpenUrl CrossRef PubMed ↵ Valtcheva , S. et al. Neural circuitry for maternal oxytocin release induced by infant cries . Nature doi: 10.1038/s41586-023-06540-4 ( 2023 ). OpenUrl CrossRef PubMed ↵ Ng , H. et al. Effects of oxytocin ablation on pup rescue, nursing behaviors and response to pup separation in early-to-mid postpartum mice . J Neuroendocrinol doi: 10.1111/jne.13247 ( 2023 ). OpenUrl CrossRef PubMed ↵ Bridges , R. S. , Mann , P. E. & Coppeta , J. S . Hypothalamic involvement in the regulation of maternal behaviour in the rat: inhibitory roles for the ventromedial hypothalamus and the dorsal/anterior hypothalamic areas . J Neuroendocrinol doi: 10.1046/j.1365-2826.1999.00322.x ( 1999 ). OpenUrl CrossRef PubMed Web of Science Hashikawa , K. et al. Esr1(+) cells in the ventromedial hypothalamus control female aggression . Nat Neurosci doi: 10.1038/nn.4644 ( 2017 ). OpenUrl CrossRef PubMed Kunwar , P. S. et al. Ventromedial hypothalamic neurons control a defensive emotion state . Elife doi: 10.7554/eLife.06633 ( 2015 ). OpenUrl CrossRef PubMed Narita , K. , Murata , T. & Matsuoka , S . The ventromedial hypothalamus oxytocin induces locomotor behavior regulated by estrogen . Physiol Behav doi: 10.1016/j.physbeh.2016.05.047 ( 2016 ). OpenUrl CrossRef PubMed ↵ Mitre , M. et al. Sex-Specific Differences in Oxytocin Receptor Expression and Function for Parental Behavior . Gend Genome 142 – 166 , doi: 10.1089/gg.2017.0017 ( 2017 ). OpenUrl CrossRef PubMed ↵ Marek , R. , Xu , L. , Sullivan , R. K. P. & Sah , P . Excitatory connections between the prelimbic and infralimbic medial prefrontal cortex show a role for the prelimbic cortex in fear extinction . Nat Neurosci doi: 10.1038/s41593-018-0137-x ( 2018 ). OpenUrl CrossRef PubMed ↵ Capuzzo , G. & Floresco , S. B . Prelimbic and Infralimbic Prefrontal Regulation of Active and Inhibitory Avoidance and Reward-Seeking . J Neurosci doi: 10.1523/JNEUROSCI.0414-20.2020 ( 2020 ). OpenUrl Abstract / FREE Full Text ↵ Sabihi , S. , Dong , S. M. , Durosko , N. E. & Leuner , B . Oxytocin in the medial prefrontal cortex regulates maternal care, maternal aggression and anxiety during the postpartum period . Front Behav Neurosci doi: 10.3389/fnbeh.2014.00258 ( 2014 ). OpenUrl CrossRef PubMed ↵ Champagne , F. , Diorio , J. , Sharma , S. & Meaney , M. J . Naturally occurring variations in maternal behavior in the rat are associated with differences in estrogen-inducible central oxytocin receptors . Proc Natl Acad Sci U S A doi: 10.1073/pnas.221224598 ( 2001 ). OpenUrl Abstract / FREE Full Text ↵ Stamatakis , A. et al. Rat dams exposed repeatedly to a daily brief separation from the pups exhibit increased maternal behavior, decreased anxiety and altered levels of receptors for estrogens (ERalpha, ERbeta), oxytocin and serotonin (5-HT1A) in their brain . Psychoneuroendocrinology doi: 10.1016/j.psyneuen.2014.11.016 ( 2015 ). OpenUrl CrossRef PubMed ↵ Li , D. et al. Involvement of Hyperpolarization-Activated Cyclic Nucleotide-Gated Channel 3 in Oxytocin Neuronal Activity in Lactating Rats With Pup Deprivation . ASN Neuro doi: 10.1177/1759091420944658 ( 2020 ). OpenUrl CrossRef PubMed ↵ Liu , X. Y. et al. Effects of Intranasal Oxytocin on Pup Deprivation-Evoked Aberrant Maternal Behavior and Hypogalactia in Rat Dams and the Underlying Mechanisms . Front Neurosci doi: 10.3389/fnins.2019.00122 ( 2019 ). OpenUrl CrossRef PubMed ↵ Wang , C. , Jiang , C. , Yuan , H. , Xiao , C. & Gao , D . Role of calbindin-D28K in estrogen treatment for Parkinson’s disease . Neural Regen Res doi: 10.3969/j.issn.1673-5374.2013.08.004 ( 2013 ). OpenUrl CrossRef PubMed Vishnyakova , P. A. et al. Expression of calbindin and calretinin in the dorsomedial and ventromedial hypothalamic nuclei during aging . Anat Rec (Hoboken ) doi: 10.1002/ar.24536 ( 2021 ). OpenUrl CrossRef PubMed ↵ Kalinowski , D. , Bogus-Nowakowska , K. , Kozlowska , A. & Rowniak , M . Expression of Calbindin, a Marker of Gamma-Aminobutyric Acid Neurons, Is Reduced in the Amygdala of Oestrogen Receptor beta-Deficient Female Mice . J Clin Med doi: 10.3390/jcm11071760 ( 2022 ). OpenUrl CrossRef ↵ Winter , J. & Jurek , B . The interplay between oxytocin and the CRF system: regulation of the stress response . Cell Tissue Res . doi: 10.1007/s00441-018-2866-2 ( 2019 ). OpenUrl CrossRef PubMed ↵ Servin-Barthet , C. et al. The transition to motherhood: linking hormones, brain and behaviour . Nat Rev Neurosci , doi: 10.1038/s41583-023-00733-6 ( 2023 ). OpenUrl CrossRef PubMed ↵ Workman , J. L. , Brummelte , S. & Galea , L. A . Postpartum corticosterone administration reduces dendritic complexity and increases the density of mushroom spines of hippocampal CA3 arbours in dams . J Neuroendocrinol doi: 10.1111/j.1365-2826.2012.02380.x ( 2013 ). OpenUrl CrossRef PubMed Leuner , B. , Fredericks , P. J. , Nealer , C. & Albin-Brooks , C . Chronic gestational stress leads to depressive-like behavior and compromises medial prefrontal cortex structure and function during the postpartum period . PLoS One doi: 10.1371/journal.pone.0089912 ( 2014 ). OpenUrl CrossRef PubMed Wonch , K. E. et al. Postpartum depression and brain response to infants: Differential amygdala response and connectivity . Soc Neurosci doi: 10.1080/17470919.2015.1131193 ( 2016 ). OpenUrl CrossRef PubMed Payne , J. L. & Maguire , J . Pathophysiological mechanisms implicated in postpartum depression . Front Neuroendocrinol doi: 10.1016/j.yfrne.2018.12.001 ( 2019 ). OpenUrl CrossRef PubMed Becker , R. O. et al. Sexual behavior and dendritic spine density of posterodorsal medial amygdala neurons in oxytocin knockout female mice . Behav Brain Res doi: 10.1016/j.bbr.2013.07.034 ( 2013 ). OpenUrl CrossRef PubMed Pekarek , B. T. , Hunt , P. J. & Arenkiel , B. R . Oxytocin and Sensory Network Plasticity . Front Neurosci doi: 10.3389/fnins.2020.00030 ( 2020 ). OpenUrl CrossRef PubMed Sequeira , M. K. & Gourley , S. L . The stressed orbitofrontal cortex . Behav Neurosci doi: 10.1037/bne0000456 ( 2021 ). OpenUrl CrossRef PubMed Radley , J. J. et al. Repeated stress induces dendritic spine loss in the rat medial prefrontal cortex . Cereb Cortex doi: 10.1093/cercor/bhi104 ( 2006 ). OpenUrl CrossRef PubMed Web of Science Mitra , R. , Jadhav , S. , McEwen , B. S. , Vyas , A. & Chattarji , S . Stress duration modulates the spatiotemporal patterns of spine formation in the basolateral amygdala . Proc Natl Acad Sci U S A doi: 10.1073/pnas.0504011102 ( 2005 ). OpenUrl Abstract / FREE Full Text Qiao , H. et al. Dendritic Spines in Depression: What We Learned from Animal Models . Neural Plast doi: 10.1155/2016/8056370 ( 2016 ). OpenUrl CrossRef PubMed ↵ Holmes , S. E. et al. Lower synaptic density is associated with depression severity and network alterations . Nat Commun doi: 10.1038/s41467-019-09562-7 ( 2019 ). OpenUrl CrossRef PubMed ↵ Borszcz , G. S . Contribution of the ventromedial hypothalamus to generation of the affective dimension of pain . Pain doi: 10.1016/j.pain.2006.02.026 ( 2006 ). OpenUrl CrossRef PubMed Wilent , W. B. et al. Induction of panic attack by stimulation of the ventromedial hypothalamus . J Neurosurg doi: 10.3171/2009.9.JNS09577 ( 2010 ). OpenUrl CrossRef PubMed Web of Science ↵ Osakada , T. et al. A dedicated hypothalamic oxytocin circuit controls aversive social learning . Nature doi: 10.1038/s41586-023-06958-w ( 2024 ). OpenUrl CrossRef PubMed ↵ Bosch , O. J . Maternal nurturing is dependent on her innate anxiety: the behavioral roles of brain oxytocin and vasopressin . Horm Behav doi: 10.1016/j.yhbeh.2010.11.012 ( 2011 ). OpenUrl CrossRef PubMed Web of Science ↵ Bosch , O. J. & Neumann , I. D . Both oxytocin and vasopressin are mediators of maternal care and aggression in rodents: from central release to sites of action . Horm Behav doi: 10.1016/j.yhbeh.2011.11.002 ( 2012 ). OpenUrl CrossRef PubMed Web of Science ↵ Neumann , I. D. , Torner , L. & Wigger , A . Brain oxytocin: differential inhibition of neuroendocrine stress responses and anxiety-related behaviour in virgin, pregnant and lactating rats . Neuroscience doi: 10.1016/s0306-4522(99)00433-9 ( 2000 ). OpenUrl CrossRef ↵ Begni , V. et al. Social isolation in rats: Effects on animal welfare and molecular markers for neuroplasticity . PLoS One doi: 10.1371/journal.pone.0240439 ( 2020 ). OpenUrl CrossRef PubMed ↵ Douglas , A. J. , Brunton , P. J. , Bosch , O. J. , Russell , J. A. & Neumann , I. D . Neuroendocrine responses to stress in mice: hyporesponsiveness in pregnancy and parturition . Endocrinology doi: 10.1210/en.2003-0461 ( 2003 ). OpenUrl CrossRef PubMed Web of Science ↵ Klampfl , S. M. & Bosch , O. J . Mom doesn’t care: When increased brain CRF system activity leads to maternal neglect in rodents . Front Neuroendocrinol doi: 10.1016/j.yfrne.2019.01.001 ( 2019 ). OpenUrl CrossRef PubMed ↵ Darnaudery , M. , Dutriez , I. , Viltart , O. , Morley-Fletcher , S. & Maccari , S . Stress during gestation induces lasting effects on emotional reactivity of the dam rat . Behav Brain Res doi: 10.1016/j.bbr.2003.12.001 ( 2004 ). OpenUrl CrossRef PubMed Web of Science ↵ Zoubovsky , S. P. et al. Chronic psychosocial stress during pregnancy affects maternal behavior and neuroendocrine function and modulates hypothalamic CRH and nuclear steroid receptor expression . Transl Psychiatry doi: 10.1038/s41398-020-0704-2 ( 2020 ). OpenUrl CrossRef PubMed ↵ Bosch , O. J. , Pohl , T. T. , Neumann , I. D. & Young , L. J . Abandoned prairie vole mothers show normal maternal care but altered emotionality: Potential influence of the brain corticotropin-releasing factor system . Behav Brain Res doi: 10.1016/j.bbr.2017.12.034 ( 2018 ). OpenUrl CrossRef PubMed ↵ Wang , T. et al. Injection of oxytocin into paraventricular nucleus reverses depressive-like behaviors in the postpartum depression rat model . Behav Brain Res doi: 10.1016/j.bbr.2017.09.012 ( 2018 ). OpenUrl CrossRef PubMed View the discussion thread. Back to top Previous Next Posted October 24, 2025. Download PDF Email Thank you for your interest in spreading the word about bioRxiv. NOTE: Your email address is requested solely to identify you as the sender of this article. Your Email * Your Name * Send To * Enter multiple addresses on separate lines or separate them with commas. You are going to email the following Long-term offspring loss in lactating rats: Neurobiological and emotional consequences in a novel animal model Message Subject (Your Name) has forwarded a page to you from bioRxiv Message Body (Your Name) thought you would like to see this page from the bioRxiv website. Your Personal Message CAPTCHA This question is for testing whether or not you are a human visitor and to prevent automated spam submissions. Share Long-term offspring loss in lactating rats: Neurobiological and emotional consequences in a novel animal model Luisa Demarchi , Alice Sanson , Anna-Lena Boos , Oliver J. Bosch bioRxiv 2025.02.17.634946; doi: https://doi.org/10.1101/2025.02.17.634946 Share This Article: Copy Citation Tools Long-term offspring loss in lactating rats: Neurobiological and emotional consequences in a novel animal model Luisa Demarchi , Alice Sanson , Anna-Lena Boos , Oliver J. Bosch bioRxiv 2025.02.17.634946; doi: https://doi.org/10.1101/2025.02.17.634946 Citation Manager Formats BibTeX Bookends EasyBib EndNote (tagged) EndNote 8 (xml) Medlars Mendeley Papers RefWorks Tagged Ref Manager RIS Zotero Tweet Widget Facebook Like Google Plus One Subject Area Neuroscience Subject Areas All Articles Animal Behavior and Cognition (7622) Biochemistry (17648) Bioengineering (13871) Bioinformatics (41880) Biophysics (21423) Cancer Biology (18558) Cell Biology (25460) Clinical Trials (138) Developmental Biology (13364) Ecology (19866) Epidemiology (2067) Evolutionary Biology (24290) Genetics (15589) Genomics (22475) Immunology (17711) Microbiology (40327) Molecular Biology (17145) Neuroscience (88473) Paleontology (666) Pathology (2827) Pharmacology and Toxicology (4816) Physiology (7635) Plant Biology (15114) Scientific Communication and Education (2044) Synthetic Biology (4286) Systems Biology (9815) Zoology (2268)