Beyond Birth Control: The Neuroscience of Hormonal Contraceptives.

Hormonal contraceptives (HCs) are one of the most highly prescribed classes of drugs in the world used for both contraceptive and noncontraceptive purposes. Despite their prevalent use, the impact of HCs on the brain remains inadequately explored. This review synthesizes recent findings on the neuroscience of HCs, with a focus on human structural neuroimaging as well as translational, nonhuman animal studies investigating the cellular, molecular, and behavioral effects of HCs. Additionally, we consider data linking HCs to mood disorders and dysregulation of the hypothalamic-pituitary-adrenal (HPA) axis and stress response as a potential mediator. The review also addresses the unique sensitivity of the adolescent brain to HCs, noting significant changes in brain structure and function when HCs are used during this developmental period. Finally, we discuss potential effects of HCs in combination with smoking-derived nicotine on outcomes of ischemic brain damage. Methodological challenges, such as the variability in HC formulations and user-specific factors, are acknowledged, emphasizing the need for precise and individualized research approaches. Overall, this review underscores the necessity for continued interdisciplinary research to elucidate the neurobiological mechanisms of HCs, aiming to optimize their use and improve women's health.

Over 300 million women globally use hormonal contraceptives (HCs; United Nations, 2019). In the United States alone, 82% of reproductive-aged women have reported using HCs at some point in their lives for both contraceptive and noncontraceptive purposes (Daniels et al., 2014). The noncontraceptive, therapeutic reasons for HC use include reducing dysmenorrhea and menorrhagia, managing acne, regulating menstrual cycles, and treating conditions such as endometriosis and polycystic ovary syndrome (Carey and Allen, 2012; Bahamondes et al., 2015). Currently, there are over 100 different formulations of HCs, which can be administered via pill (i.e., oral contraceptives, OCs), through long-acting reversible contraceptives (LARCs) like hormonal intrauterine devices (IUDs) and contraceptive implants, transdermal patches, or by injection (e.g., Depo-Provera). Each type of HC relies on synthetic hormones—either a synthetic progestin only or a synthetic progestin in combination with a synthetic estrogen. The estrogen component of combined HCs is typically ethinyl estradiol (EE), while the progestin component varies across several types and classes (Darney, 1995; Stanczyk et al., 2013; Hampson, 2023; Pletzer et al., 2023). HCs achieve contraception primarily through negative feedback on the hypothalamic-pituitary-gonadal (HPG) axis (Fig. 1; Davis and Hackney, 2017). By doing so, many HCs diminish the production of endogenous estradiol and progesterone by the ovaries, suppress cyclical hormonal fluctuations, and inhibit ovulation (Fleischman et al., 2010; Hampson, 2023). Thus, HCs are major modifiers of the hormonal milieu for many women, often across years of their lives. It should be noted, however, that not all forms of HCs work by inhibiting the HPG axis. For example, hormonal IUDs achieve their contraceptive effects primarily by altering the local endometrial environment (i.e., thinning of the endometrial lining and thickening of cervical mucus; Rivera et al., 1999; Smith-McCune et al., 2020). Beyond their effects on the HPG axis, relatively little is known about the impact of HCs on the brain. HCs have been available for over 60 years (Christin-Maitre, 2013), and given the large body of research showing that the nervous system is a major target for the actions of endogenous ovarian hormones (McEwen, 2020; Rehbein et al., 2021), there is a strong need for more neuroscience studies focused on the potential consequences of disruptions to the hormonal milieu due to HC use (Petersen et al., 2023). Recent efforts have gradually begun to address this knowledge gap, identifying the neural and functional effects of HCs. The emerging field of HC neuroscience has been advanced in various subfields of research including those discussed in our 2024 Society for Neuroscience Mini-symposium. These include human neuroimaging (Heller et al., 2022) as well as translational, nonhuman animal studies investigating the cellular, molecular, and behavioral effects of HCs (Lacasse et al., 2022; Porcu et al., 2019). This work, along with preclinical and clinical research aimed at understanding how HCs influence risk and resilience to central nervous system (CNS) conditions such as stroke (Reddy et al., 2022) and mood disorders (Tronson and Schuh, 2022; Johansson et al., 2023), suggests that the brain is indeed sensitive to HCs and that HC use has significant implications for health and disease (other areas of HC research not considered here are also consistent with this idea including effects of HCs on human cognition and age-related cognitive impairment; see Beltz et al., 2015; Karim et al., 2016; Hampson et al., 2022; Gregory et al., 2023; Gurvich et al., 2023). In this review, we present recent research from each of these areas and discuss other novel avenues of investigation in this nascent field, including the effects of HCs on the adolescent brain. We conclude by discussing important themes that emerge from these studies, emphasizing that the influence of HCs on the brain and behavior is often nuanced and suggest promising areas of focus for future work. Our goal is that with growing interest in studying factors influencing women's health (Deems and Leuner, 2020; Taylor et al., 2021; Galea and Parekh, 2023), greater attention will be given to the neuroscientific study of HCs. HC Effects on the Human Brain: Evidence from Structural Neuroimaging Neuroimaging studies have started to characterize the effects of HCs on the CNS (Pletzer and Kerschbaum, 2014; Montoya and Bos, 2017; Beltz and Moser, 2020; Taylor et al., 2021). In addition to functional connectivity changes associated with HC use (Brønnick et al., 2020), there is a growing body of research dedicated to exploring the structural changes that occur during HC use compared with the natural menstrual cycle (Heller et al., 2022). These structural neuroimaging studies have revealed a nuanced landscape of changes throughout the brain (Fig. 2), including both increases and decreases in volume in various cortical and subcortical gray matter brain regions mediating affective and cognitive processing as well as changes in cortical thickness (Heller et al., 2022; Rehbein et al., 2021). Decreases in cortical volumes in women using HCs compared with naturally cycling controls have been observed in several brain regions including the bilateral middle frontal gyrus, the superior frontal gyrus (Pletzer et al., 2015), the parahippocampal gyrus (Lisofsky et al., 2016), the right anterior cingulate gyrus, the right lingual gyrus (De Bondt et al., 2016), and the left fusiform gyrus (De Bondt et al., 2013). Subcortically, volume decreases have been reported in the right putamen (Sharma et al., 2020b), the left hippocampus (Pletzer et al., 2019), and left amygdala (Lisofsky et al., 2016). Moreover, decreased gray matter volume has been reported in the right hemisphere of the cerebellum (De Bondt et al., 2016). In contrast, increases in cortical volumes have also been noted in various regions throughout the brain. These include structures in the bilateral prefrontal (Pletzer et al., 2010; De Bondt et al., 2013; Sharma et al., 2020b) and temporal cortex (Pletzer et al., 2010), as well as the bilateral postcentral gyrus, the supramarginal gyrus, the inferior occipital cortex, the cerebellum, the right middle occipital cortex, and the right inferior parietal gyrus (Pletzer et al., 2010). Further, HC use has been associated with localized decreases in cortical thickness (i.e., average distance between gray matter surface of the cerebral cortex and the underlying white matter) in the left anterior and posterior cingulate gyrus, the left insula, as well as the bilateral pars triangularis and orbitofrontal gyrus (Petersen et al., 2015). As of now, there is no clear pattern emerging from these findings, highlighting the complexity of structural brain changes associated with HC use (for review see Heller et al., 2022) and the need for more work to determine the causes of variability across studies. While structural neuroimaging has identified volume and cortical thickness changes in women using HCs, it falls short of understanding the underlying neurobiological mechanisms. Are changes in volume and cortical thickness indicative of alterations in neuronal and synaptic density within the cortical layers, or do they reflect changes in factors such as blood flow and water content? To shed light on possible microscopic phenomena, diffusion magnetic resonance imaging (dMRI) has emerged as a promising avenue of exploration (Novikov et al., 2019; Jelescu et al., 2020). By probing water molecule diffusion within neural tissues (Basser et al., 1994), dMRI can uncover subtle changes in microarchitecture, yielding information on neuronal density, axonal integrity, myelination, or dendritic morphology in vivo (Beaulieu, 2002; S. K. Song et al., 2002; Alexander et al., 2007). In two cross-sectional dMRI studies, increased fractional anisotropy (FA) and mean diffusivity (MD) were reported in HC users (DeBondt et al., 2013; Sharma et al., 2020b). FA indicates the degree of diffusion anisotropy, reflecting how much water molecules preferentially move along the orientation of fiber tracts versus perpendicular to them. Higher FA values suggest greater directionality of water diffusion, typically indicating more organized and intact white matter fiber tracts. On the other hand, MD gives a broader measure of the magnitude of water diffusion, reflecting tissue properties such as cellularity, membrane integrity, and extracellular space. Increased MD often indicates greater water diffusion, which can be associated with various changes such as cellular swelling, reduced tissue density, or damage to cellular structures (Beaulieu, 2014). Together, the available evidence from dMRI points to HCs having microstructural effects, but with only two published studies, it remains unclear whether these findings can be generalized to the broader population. HC Effects on the Brain: Animal Models Animal models of HC exposure complement human studies. Although human studies show that HCs affect the brain and behavior (Pletzer and Kerschbaum, 2014; Montoya and Bos, 2017; Beltz and Moser, 2020; Taylor et al., 2021; Heller et al., 2022; J. Y. Song et al., 2023), it is impossible to ethically and effectively study the neurobiological mechanisms underlying these effects at a cellular and molecular level of analysis. Additionally, human experiments are complicated by individual differences in genetics and experiences, as well as variables related to HCs themselves, such as hormone formulation, dose, and schedule (Beltz, 2022; Hill and Mengelkoch, 2023). Animal models of HC exposure thus provide an important way to address these limitations of human-subject studies, as they offer the ability to control and experimentally manipulate these factors (Hilz, 2022; Lacasse et al., 2022; Tronson and Schuh, 2022). Most commonly, rodent models of HCs employ administration of EE and the progestin levonorgestrel (LNG) to intact adult female rodents (for a review on animal models of HCs, see Hilz et al., 2022; Lacasse et al., 2022). Like humans, administration of HCs to rodents leads to HPG axis suppression, acyclicity, and anovulation (Kuhl et al., 1984; Porcu et al., 2012; Graham and Milad, 2013; Santoru et al., 2014; Simone et al., 2015; Lacasse et al., 2022). Research using nonhuman animal models have begun to provide valuable insights into the neurobiological processes impacted by contraceptive hormones. For example, rodent studies suggest that HCs decrease cerebrocortical neuroactive steroid levels, including progesterone and the progesterone metabolite allopregnanolone (Porcu et al., 2019). Various neurotransmitter systems have also been shown to be altered by HCs. For example, Jori and Dolfini (1976) demonstrated that administration of mestranol (a pro-drug for EE), in combination with various progestins, markedly reduced concentrations of striatal dopamine in female rats while Hilz et al. (2022) found that LNG reduced dopamine cell number in the substantia nigra. Rodent studies have additionally shown effects of HCs on the serotonin system (Baker et al., 1977; Daabees et al., 1981; Vega-Rivera et al., 2013), cholinergic (Mennenga et al., 2015) and noradrenergic systems (Simone et al., 2015), as well as glutamatergic and GABAergic systems (Daabees et al., 1981; Follesa et al., 2002; Porcu et al., 2012). Of particular interest in many rodent neurobehavioral studies are the effects of HCs related to hippocampal neuroplasticity. The hippocampus, a key brain region for cognition and spatial memory, is critically involved in conditions like major depressive disorder (Campbell and MacQueen, 2004; MacQueen and Frodl, 2011) and Alzheimer's disease (Knopman et al., 2021). It is rich in receptors for gonadal hormones such as estrogens (Almey et al., 2015), progesterone (Brinton et al., 2008; Thomas and Pang, 2012), and androgens (Sarkey et al., 2008; Moghadami et al., 2016) and is significantly influenced by ovarian hormones like 17β-estradiol (E2). Animal studies demonstrate that E2 enhances spatial memory (Frick et al., 2018), synaptic plasticity (Luine and Frankfurt, 2013; Finney et al., 2020), and neurogenesis (Yagi et al., 2022, 2023) in the hippocampus. Thus, the hippocampus is a key region for studying the brain-based effects of HCs. Rodent studies have investigated how contraceptive hormones like EE and various progestins influence hippocampus-dependent spatial tasks, with resulting effects often dependent on dose and specific hormone administered. For example, Simone et al. (2015) found that EE and/or LNG impaired recognition memory in the novel object recognition test and that EE administered at higher doses enhanced novel object preference compared with vehicle-treated female rats. In addition, Lacasse et al. (2023) demonstrated that in female rats, LNG alone promoted hippocampus-mediated place memory in the dual-solution plus maze task. However, the same dose of LNG when combined with EE shifted memory use from hippocampus-mediated place memory to striatum-mediated response memory suggesting that effects of HCs administered in combination are not always the same as their effects in isolation. It should be noted that other rodent studies did not find effects of contraceptive hormones on hippocampal spatial memory tasks (Braden et al., 2011; Santoru et al., 2014; Boi et al., 2022), discrepancies which may be due to variations in experimental conditions and the specific memory domains examined. The neurobiological mechanisms underlying HC-induced changes in cognitive performance are still unknown. One candidate is brain-derived neurotrophic factor (BDNF). BDNF is strongly influenced by E2 and progesterone (Concas et al., 2022) and is known to play a crucial role in learning and memory processes by promoting neuronal survival, growth, and synaptic plasticity within the hippocampus (Leal et al., 2015; Kowiański et al., 2018). Given these links, Simone et al. (2015) investigated BDNF mRNA in the hippocampus of female rats chronically treated with the combination of EE and LNG and found reduced expression, particularly in the dentate gyrus and CA regions. Hippocampal BDNF protein levels were also reduced when higher doses of EE and LNG were administered. Similarly, Boi et al. (2022) reported that chronic treatment with EE and LNG significantly decreased hippocampal BDNF protein levels after treatment discontinuation; however, in this study BDNF mRNA expression remained unchanged. Hippocampal neurogenesis, the generation of new neurons in the hippocampus, is essential for brain plasticity and function of this region (Leuner and Gould, 2010). Hippocampal neurogenesis is also sensitive to hormones (Hodges et al., 2022), including contraceptive hormones. In a study by Liu et al. (2010), the effects of synthetic progestins, such as LNG and medroxyprogesterone acetate (MPA), on neurogenesis were examined. They found that LNG promoted the growth of rat neural progenitor cells in vitro but did not significantly boost new cell growth in the female rat hippocampus in vivo. MPA, on the other hand, inhibited neural progenitor cell growth in vitro and had no notable impact on neurogenesis in vivo. However, when combined with E2, both progestins increased cell proliferation but also elevated apoptosis, indicating a detrimental impact on cell survival despite enhanced proliferation. Thus, type of progestin and whether or not it is combined with estrogen can have differential effects on neurogenesis. Overall, rodent studies have demonstrated that the hippocampus is sensitive to the actions of contraceptive hormones. The evidence from spatial memory tasks, BDNF, and neurogenesis once again highlights that HCs have nuanced effects which are often complex and thus warrant greater investigation. HC Effects on Mood and the Hypothalamic-Pituitary-Adrenal (HPA) Axis Potential links between the use of HCs and mood disturbance, particularly depressive changes, is mixed. Whereas some studies show no effect of HCs on mood, others demonstrate that HCs can improve mood for some users (Keyes et al., 2013; Schaffir et al., 2016; Lundin et al., 2022; Kraft et al., 2024). HCs have also been shown to increase the risk of depression in a subset of people (Poromaa and Segebladh, 2012; Skovlund et al., 2016; Lewis et al., 2019). Indeed, several large-scale epidemiological studies have linked HC use to risk for depression (Skovlund et al., 2016; Zettermark et al., 2018; Johansson et al., 2023), making the exploration of potential mediators underlying susceptibility to depression in HC users a focal point of HC research. One potential explanation for the mood effects of HCs is via regulation of the hypothalamic-pituitary-adrenal (HPA) axis and altered stress responsiveness that occurs with HC use. Indeed, a highly consistent finding is the observation that HC use blunts cortisol reactivity (Kirschbaum et al., 1995; Boisseau et al., 2013; Hertel et al., 2017; Mordecai et al., 2017; Gervasio et al., 2022). In some studies, HCs have also been shown to increase basal levels of glucocorticoids (Hertel et al., 2017; Lovallo et al., 2019). Notably, blunted cortisol reactivity is a risk factor for depression (Luby et al., 2003; Pruessner et al., 2003; Cunningham et al., 2021), while elevated baseline cortisol may indicate a chronic stress-like phenotype, which also enhances depression risk (Baumeister et al., 2016). How HCs modulate the HPA axis is not known. Animal models are well suited to extending observations from human studies to identify specific cellular, molecular, and neuroendocrine mechanisms by which HCs impact HPA axis function (Tronson and Schuh, 2022). Toward this end, Schuh et al. (2024) recently created and validated a mouse model of OC exposure to mimic the administration and timing of human OC use. This protocol successfully replicates key outcomes observed in human OC users, including disrupted estrous cycles and a blunted stress response, demonstrating the model's translational utility (Schuh et al., 2024). OC-treated mice also exhibited a disruption in sucrose preference, an anhedonia-like effect, without broad changes in anxiety or stress-coping behaviors (Schuh et al., 2024). This work suggests that OC exposure affects specific psychological and physiological processes rather than causing generalized depressive-like behavior. Additional research in both animals and humans is needed to better understand the mechanisms underlying HPA axis disruption with HC use and whether HC effects on the stress response drive mood-related HC side effects. Further, because not all individuals who use HCs experience adverse effects on mood, it is crucial to better understand what contributes to individual differences in resilience versus susceptibility (Tronson and Schuh, 2022). It may be that increased depression susceptibility is not a general feature of HCs, but only of certain formulations (Schaffir et al., 2016; Rapkin et al., 2019, Bürger et al., 2021). Increased risk could also be associated with person-specific variables such as possessing specific genetic variants (Hamstra et al., 2015) or a prior history of depression (Bengtsdotter et al., 2018). Animal models can be used to isolate these candidate risk factors to identify whether and how each factor contributes to risk for HC-triggered depression. Adolescence as a Unique Period of Sensitivity to HCs Age at first use is an important factor to consider when investigating the effects of HCs on the brain and behavior. HCs are approved for use after menarche and it has been estimated that ~1 in 4 adolescent females will be exposed to HCs by the age of 18 (Bonny et al., 2015; Abma and Martinez, 2023). Despite the high prevalence of HC use among teens, almost all neuroscience studies to date have been done in adult HC users or, for preclinical studies, in rodent models using adult females. There is growing recognition to consider that since the adolescent brain is not fully mature, it may be uniquely sensitive to the neurobehavioral effects of HCs (Cahill, 2018; Brønnick et al., 2020). Research addressing this possibility is critical as a growing number of studies suggest that use of HCs, specifically among adolescents, enhances risk for major depression including years later in adulthood, even after cessation of use (Anderl et al., 2020, 2022). Accordingly, adolescents who use HCs are also more likely to also use antidepressants (Lindberg et al., 2012; Skovlund et al., 2016) and are at greater risk for suicide (Skovlund et al., 2018; de Wit et al., 2020). Maturation of the adolescent brain involves changes to gray and white matter and is shaped by processes such as synaptic pruning and myelination that reconfigure brain connectivity into the adult form (Ladouceur et al., 2012; Spear, 2013). Evidence from both animals and humans indicates that for females, various aspects of adolescent brain development require ovarian hormones (Jursaka et al., 2013; Koolschijn, et al., 2014; Herting and Sowell, 2017; Delevich et al., 2021). Indeed, adolescence is widely regarded to be a sensitive period marked by the activation of the HPG axis and the production of gonadal hormones which have long-lasting “organizational”? effects in sculpting neural circuits to influence behavior (Schulz and Sisk, 2016). Thus, it stands to reason that, through suppression of endogenous hormones and/or the introduction of exogenous synthetic hormones, HCs in adolescence have the potential to impact hormone-mediated neurodevelopmental processes. Neuroimaging studies from the Ismail lab have examined pubertal-onset use of OCs and found that it results in greater white matter volume compared with adult-onset OC use in brain structures (e.g., hippocampus, fusiform gyrus, and precuneus) implicated in emotional regulation and memory, possibly suggesting greater myelination (Sharma et al., 2020a). Functional differences in the brains of those who began taking OCs during adolescence that are not observed in those who began taking OC during adulthood have also been reported (Marecková et al., 2014; Sharma et al., 2020a,b). For example, Sharma et al. (2020b) showed that OC use during puberty/adolescence alters brain activation during working memory processing. In addition, OC use has been shown to modify resting state functional connectivity (Sharma et al., 2020a) such that those who began taking OCs during early adolescence display greater resting state functional connectivity compared with adult-onset OC users, specifically in the salience network which is known to link cognition with emotion/interoception. Thus, although findings are limited, they nonetheless demonstrate that OCs alter brain structure, function, and resting state functional connectivity during adolescence. Animal models can also be leveraged to understand the effects of HC exposure in adolescence and can provide insights at a cellular and molecular level of resolution not feasible in humans. This is an active area of investigation in the Leuner laboratory which studies the effects of adolescent exposure to the contraceptive hormones EE and LNG on the medial prefrontal cortex. Like the human prefrontal cortex (Koolschijn, et al., 2014; Herting and Sowell, 2017; Chini and Hanganu-Opatz, 2021), the rodent medial prefrontal cortex is a late developing, hormone-sensitive brain region that mediates many emotional, cognitive, and social behaviors which mature during adolescence (Shaw et al., 2020; Delevich et al., 2021). Unpublished data from the Leuner lab suggests that HCs can impact molecular and functional markers of cortical maturation in female rats, as well as neuroimmune factors which regulate these processes. The extent to which the effects of HCs on the adolescent brain point to shifts in typical developmental trajectories and whether they are permanent remains to be determined. As discussed above, HC use has been associated with HPA axis dysregulation, and new data suggests that this is likely true for adolescence as well. For example, work from the Leuner lab has shown that female rats treated with EE and LNG in adolescence have higher baseline corticosterone as well as a blunted corticosterone response to acute stress. Recent unpublished data by Lacasse et al. further highlight the increased sensitivity of adolescent female rats to the HPA axis-disrupting effects of HCs compared with adult females. Specifically, both adolescents and adults exhibit an attenuated corticosterone response to restraint stress when administered EE and LNG, but this dampening of corticosterone levels is significantly more pronounced in adolescents. Notably, these rodent data align with work by Sharma et al. (2020b) which found that OC use during early adolescence gives rise to a blunted cortisol response following stress exposure compared with females that began taking OC in adulthood. Whether these features of HPA dysregulation contribute to increased depression susceptibility in adolescent HC users is an open question. Overall, the effects of contraceptive hormone use during adolescence are just beginning to be identified. Emerging evidence indicates effects of adolescent HCs on the brain and HPA axis and further underscore the importance of considering age as a critical variable in HC research. HCs and CNS Injury Animal models have also significantly contributed to our understanding of how HC use contributes to outcomes following CNS injury like cerebral ischemia that could be a primary or secondary consequence of stroke or traumatic injury, respectively. HCs, specifically OCs, are one of the causes of increased stroke risk among young women (Leppert et al., 2020; Johansson et al., 2022). The association between OC use and increased risk and severity of stroke was first established in the 1960s when a woman on OCs died from coronary thrombosis within a year after the US Food and Drug Administration-approved OCs (Boyce et al., 1963; Tyler, 1963; Mann et al., 1975; Stadel, 1981; Sartwell and Stolley, 1982; Thorogood and Vessey, 1990; Rosendaal, 1997; Tanis et al., 2001; Tanis and Rosendaal, 2003). More recent data suggests that OC use may increase the risk of stroke, especially during the first year of use, possibly due to immediate changes in hemostatic balance (Johansson et al., 2022). In addition, long-term OC exposure is associated with increased blood clotting, plasma fibrinogen, coagulation factors, and total cholesterol levels, contributing to venous and arterial thrombosis (Bloemenkamp et al., 1995; Jick et al., 1995; Spitzer et al., 1996; Morimont et al., 2021; Teal and Edelman, 2001). It is also known that combining OC use with tobacco smoking magnifies risk for and severity of stroke in women as compared with women who use OC and do not smoke (Goldbaum et al., 1987; Hannaford, 2000; Keeling, 2003; Goldstein et al., 2006; Bhat et al., 2008). Since women have higher lifetime risk of stroke, higher mortality from stroke, and a higher tendency for recurrent strokes than men, a better understanding of short- and long-term exposure to OCs in the context of preventable risk factors like smoking is needed to mitigate the risk and severity of cerebral ischemia (Girijala et al., 2017). Employing a female rat model of global and focal cerebral ischemia, work from the Raval lab has found that as little as 2 weeks of combined exposure to cigarette smoking-derived nicotine and OC (using dosing regimens similar to women's OC usage) resulted in severe postischemic hypoperfusion and brain damage (Raval et al., 2011; D’Adesky et al., 2021; Patel et al., 2022). Further, studies indicate that OCs exacerbate nicotine toxicity by altering mitochondrial function, particularly through defects in the terminal enzyme of the electron transport chain, cytochrome c oxidase (Raval et al., 2012; Diaz and Raval, 2021). These mitochondrial defects alter energy metabolism in the brains of female rats exposed simultaneously to both OC and nicotine (Diaz and Raval, 2021). Specifically, nicotine and OC exposure increased brain glycolysis in an age-dependent manner. Since glucose metabolism is critical for brain physiology, altered glycolysis deteriorates neural function, thereby exacerbating ischemic brain damage. The observed defects in energy metabolism and severity of ischemic damage due to combination of nicotine and OC were more pronounced in adolescent rats as compared with adult rats, suggesting that adolescent brains are more sensitive to OCs as presented in the previous section (Diaz and Raval, 2021). Overall, these findings necessitate further investigation to determine the ideal dose or combination of OCs that can reduce the severity of CNS injury while maintaining contraceptive and noncontraceptives benefits. Similar to the aforementioned effects of HCs on mood, they highlight the importance of weighing risks and benefits critically for each individual prior to HC or use. Methodological Challenges in HC Neuroscience Research and Future Directions The field of HC neuroscience is new, and as in any new field, there are challenges to overcome (Beltz, 2022; Petersen et al., 2023). The synthetic hormones in HCs bind to endogenous steroid hormone receptors, affecting brain regions and neural circuits sensitive to these hormones, while simultaneously suppressing endogenous steroid hormone production and preventing their cyclical release (Lacasse et al., 2022; Hampson, 2023). Preclinical data also suggest that HCs affect steroid hormones in the brain (Porcu et al., 2019). Thus, disentangling the effects of endogenous hormonal alterations, whether peripherally and/or centrally, from those related to synthetic hormone agonist effects is a significant methodological hurdle. The pharmacological nuances of HCs pose further obstacles. HC formulations vary in chemical composition, dosage, and route of administration (Hampson, 2023). Although most OCs contain both synthetic estrogen and progestin, the estrogen–progestin ratio varies significantly among formulations (Dickey, 2021). Further, some oral and many non-oral forms (e.g., IUDs, injection) contain only progestin. The progestin component of contraceptives also varies across several kinds and classes of progestin, each with its own specific profile of pharmacological efficacy, kinetics, and off-target effects including varying degrees of androgenicity (Phillips et al., 1990; Darney, 1995; Fuhrmann et al., 1996; Schindler et al., 2003; Kuhl, 2005; Sitruk-Ware and Nath, 2010; Africander et al., 2011; Giatti et al., 2016; Pletzer et al., 2023). For OCs, hormonal regimens also vary based on whether the drug dose is constant or variable across a pill pack (e.g., monophasic vs multiphasic doses). This diversity in HCs makes comprehensive study challenging. Nonetheless, systematically examining these variations in HCs is crucial as they likely exert different effects on neurobiological systems. Several user-specific variables also pose methodological challenges, as they may modify HC effects on the brain. These include the age at which HC use begins (Cahill, 2018; Kheloui et al., 2023), the duration of use (Pletzer et al., 2019; Davignon et al., 2024), and individual genetic differences (Hamstra et al., 2015; Gravelsins et al., 2021; Klump and Di Dio, 2022). Thus, studying the neurobiological effects of HCs requires considering the specific HC formulation and the characteristics of the user, rather than simply categorizing individuals as HC users or nonusers. The future success of HC neuroscience will require integrating the nuances of HC use into studies while simultaneously harnessing the latest methodological and technical approaches. For example, newer neuroimaging studies aim to go beyond cross-sectional designs by implementing a dense-sampling methodology and in doing so, build upon a deep-phenotyping approach that tracks individuals over extended periods ranging from days to months (Poldrack et al., 2015; Barth et al., 2016; Gordon et al., 2017; Pritschet et al., 2020, 2021; Taylor et al., 2020; Grotzinger et al., 2024; Heller et al., 2024). This approach has been applied in recent work by Heller et al. in which a participant underwent extensive brain scans and blood draws for five consecutive weeks on three separate assessment periods within one year, resulting in a total of 75 scans and blood draws. One assessment period took place during combined HC use, while the other two occurred during the natural menstrual cycle—before and after initiating the contraceptive regimen. With the dense sampling approach, it is thus possible to capture subtle changes overlooked in less frequent sampling and has the added benefit of being able to directly compare naturally cycling and hormonally suppressed states within the same individual. Combining sophisticated neuroimaging techniques with longitudinal designs are needed to identify effects of HCs over time, at different life stages, and after discontinuation of use in order to bridge the existing knowledge gaps. Addressing some of the outstanding questions about the impact of HCs on the brain may be more accessible in animal models, and as such, direct translation between human and animal HC studies are a vital next step. For example, neuroimaging methods that are used in clinical settings can also be used in animals (Heller et al., 2022; J. Y. Song et al., 2023). Importantly, studies using animal models can go beyond documenting HC effects in the CNS and employ targeted molecular and biochemical tools, as well as pharmacological and genetic manipulations, to identify causal mechanisms. Finally, there is growing interest in preclinical and clinical HC research to examine the interactions between HC use and other systems such as the immune system (Mengelkoch et al., 2024) and gut microbiome (Kheloui et al., 2023). Integrated translational work in animal models and humans provides opportunities for critical new insights into the impact of HC use on the nervous system (see Taylor et al., 2021; Beltz, 2022; J. Y. Song et al., 2023; Hill and Mengelkoch, 2023 for additional recommendations for research approaches to study HCs).

The emerging neuroscience of HCs highlights the profound influence these pharmaceuticals exert on the brain. While significant progress has been made, the current body of research indicates that we are only beginning to explore this area in depth. The intricate interplay between synthetic hormones and brain function requires a more nuanced approach to identify the differential effects of multiple formulations of HCs, their varying pharmacological properties, and individual user characteristics. The methodological challenges and varied findings highlight the need for continued interdisciplinary research to identify how and where HCs act in the brain and the short -and long-term implications of HC use. Here, we have summarized the current knowledge, delineated the next steps in our research trajectories, and highlighted key research gaps. The overarching takeaway from this exploration is clear: while our understanding of HCs and their impact on the brain is growing, there is still an enormous amount of work needed to fully understanding the extent of their effects and optimize their use for women's health. Footnotes This work was supported by the Centre for Addiction and Mental Health Discovery Fund to J.M.L.; German Research Foundation Grant #544183227 to C.H.; Natural Sciences and Engineering Council of Canada Discovery Grant #2020-04302 to N.I.; Veterans Affairs #RX003506 and American Heart Association #23TPA1142407 to A.P.R.; Oscar Stern Award, UM Eisenberg Family Depression Center to N.C.T.; Eunice Kennendy Shriver National Institute for Child Health and Human Development R21-HD109618 to B.L. The authors declare no competing financial interests. - Correspondence should be addressed to Benedetta Leuner at leuner.1{at}osu.edu.