Palatable food reduces anxiety-like behaviors and HPA axis responses to stress in female rats in an estrous-cycle specific manner.

Eating tasty foods dampens responses to stress - an idea reflected in the colloquial term 'comfort foods'. To study the neurobiological mechanisms by which palatable foods provide stress relief, we previously characterized a limited sucrose intake (LSI) paradigm in which male rats are given twice-daily access to 4 ml of 30% sucrose solution (vs. water as a control), and subsequently have reduced hypothalamic-pituitary-adrenocortical (HPA) axis responsivity and anxiety-related behaviors. Notably, women may be more prone to 'comfort feeding' than men, and this may vary across the menstrual cycle, suggesting the potential for important sex and estrous cycle differences. In support of this idea, LSI reduces HPA axis responses in female rats during the proestrus/estrus (P/E), as opposed to the diestrus 1/diestrus 2 (D1/D2) estrous cycle stage. However, the effect of LSI on anxiety-related behaviors in females remains unknown. Here we show that LSI reduced stress-related behaviors in female rats in the elevated plus-maze and restraint tests, but not in the open field test, though only during P/E. LSI also decreased the HPA axis stress response primarily during P/E, consistent with prior findings. Finally, cFos immunolabeling (a marker of neuronal activation) revealed that LSI increased post-restraint cFos in the central amygdala medial subdivision (CeM) and the bed nucleus of the stria terminalis posterior subnuclei (BSTp) exclusively during P/E. These results suggest that in female rats, palatable food reduces both behavioral and neuroendocrine stress responses in an estrous cycle-dependent manner, and the CeM and BSTp are implicated as potential mediators of these effects.

Palatable Food Affects HPA Axis Responsivity and Forebrain Neurocircuitry in an Estrous Cycle-specific Manner in Female Rats.

Eating palatable foods can provide stress relief, but the mechanisms by which this occurs are unclear. We previously characterized a limited sucrose intake (LSI) paradigm in which twice-daily access to a small amount of 30% sucrose (vs. water as a control) reduces hypothalamic-pituitary-adrenocortical (HPA) axis responses to stress and alters neuronal activation in stress-regulatory brain regions in male rats. However, women may be more prone to 'comfort feeding' behaviors than men, and stress-related eating may vary across the menstrual cycle. This suggests that LSI effects may be sex- and estrous cycle-dependent. The present study therefore investigated the effects of LSI on HPA axis stress responsivity, as well as markers of neuronal activation/plasticity in stress- and reward-related neurocircuitry in female rats across the estrous cycle. We found that LSI reduced post-restraint stress plasma ACTH in female rats specifically during proestrus/estrus (P/E). LSI also increased basal (non-stress) FosB/deltaFosB- and pCREB-immunolabeling in the basolateral amygdala (BLA) and central amygdala specifically during P/E. Finally, Bayesian network modeling of the FosB/deltaFosB and pCREB expression data identified a neurocircuit that includes the BLA, nucleus accumbens, prefrontal cortex, and bed nucleus of the stria terminalis as likely being modified by LSI during P/E. When considered in the context of our prior results, the present findings suggest that palatable food reduces stress responses in female rats similar to males, but in an estrous cycle-dependent manner. Further, the BLA may contribute to the LSI effects in both sexes, whereas the involvement of other brain regions appears to be sex-dependent.

Sucrose-induced plasticity in the basolateral amygdala in a 'comfort' feeding paradigm.

A history of intermittent, limited sucrose intake (LSI) attenuates the hypothalamic-pituitary-adrenocortical (HPA) axis stress response, and neuronal activity in the basolateral amygdala (BLA) is necessary for this HPA-dampening. LSI increases the expression of plasticity-associated genes in the BLA; however, the nature of this plasticity is unknown. As BLA principal neuron activity normally promotes HPA responses, the present study tests the hypothesis that LSI decreases stress-excitatory BLA output by decreasing glutamatergic and/or increasing GABAergic inputs to BLA principal neurons. Male rats with unlimited access to chow and water were given additional access to 4 ml of sucrose (30%) or water twice daily for 14 days, and BLA structural and functional plasticity was assessed by quantitative dual immunolabeling and whole-cell recordings in brain slices. LSI increased vesicular glutamate transporter 1-positive (glutamatergic) appositions onto parvalbumin-positive inhibitory interneurons, and this was accompanied by increased expression of pCREB, a marker of neuronal activation that is mechanistically linked with plasticity, within parvalbumin interneurons. LSI also increased the paired-pulse facilitation of excitatory, but not inhibitory synaptic inputs to BLA principal neurons, without affecting postsynaptic excitatory or miniature excitatory and inhibitory postsynaptic currents, suggesting a targeted decrease in the probability of evoked synaptic excitation onto these neurons. Collectively, these results suggest that LSI decreases BLA principal neuron output by increasing the excitatory drive to parvalbumin inhibitory interneurons, and decreasing the probability of evoked presynaptic glutamate release onto principal neurons. Our data further imply that palatable food consumption blunts HPA stress responses by decreasing the excitation-inhibition balance and attenuating BLA output.

HPA Axis Interactions with Behavioral Systems.

Perhaps the most salient behaviors that individuals engage in involve the avoidance of aversive experiences and the pursuit of pleasurable experiences. Engagement in these behaviors is regulated to a significant extent by an individual's hormonal milieu. For example, glucocorticoid hormones are produced by the hypothalamic-pituitary-adrenocortical (HPA) axis, and influence most aspects of behavior. In turn, many behaviors can influence HPA axis activity. These bidirectional interactions not only coordinate an individual's physiological and behavioral states to each other, but can also tune them to environmental conditions thereby optimizing survival. The present review details the influence of the HPA axis on many types of behavior, including appetitively-motivated behaviors (e.g., food intake and drug use), aversively-motivated behaviors (e.g., anxiety-related and depressive-like) and cognitive behaviors (e.g., learning and memory). Conversely, the manuscript also describes how engaging in various behaviors influences HPA axis activity. Our current understanding of the neuronal and/or hormonal mechanisms that underlie these interactions is also summarized. © 2016 American Physiological Society. Compr Physiol 6:1897-1934, 2016.

Activation of physiological stress responses by a natural reward: Novel vs. repeated sucrose intake.

Pharmacological rewards, such as drugs of abuse, evoke physiological stress responses, including increased heart rate and blood pressure, and activation of the hypothalamic-pituitary-adrenal (HPA) axis. It is not clear to what extent the natural reward of palatable foods elicits similar physiological responses. In order to address this question, HPA axis hormones, heart rate, blood pressure and brain pCREB immunolabeling were assessed following novel and repeated sucrose exposure. Briefly, adult, male rats with ad libitum food and water were given either a single (day 1) or repeated (twice-daily for 14 days) brief (up to 30 min) exposure to a second drink bottle containing 4 ml of 30% sucrose drink vs. water (as a control for bottle presentation). Sucrose-fed rats drank more than water-fed on all days of exposure, as expected. On day 1 of exposure, heart rate, blood pressure, plasma corticosterone, and locomotion were markedly increased by presentation of the second drink bottle regardless of drink type. After repeated exposure (day 14), these responses habituated to similar extents regardless of drink type and pCREB immunolabeling in the hypothalamic paraventricular nucleus (PVN) also did not vary with drink type, whereas basolateral amygdala pCREB was increased by sucrose intake. Taken together, these data suggest that while sucrose is highly palatable, physiological stress responses were evoked principally by the drink presentation itself (e.g., an unfamiliar intervention by the investigators), as opposed to the palatability of the offered drink.

Metabolic consequences of chronic intermittent mild stress exposure.

Chronic stress in humans has divergent effects on food intake, with some individuals reporting increased vs. decreased food intake during stress. This divergence may depend in part on stress intensity, with higher-intensity stressors preferentially promoting anorexia. Consistent with this idea, rodents given a high-intensity chronic variable stress paradigm have robustly decreased food intake and body weight gain. However, the metabolic effects of a less intense chronic stress paradigm are not clear. Thus in the present study, adult male rats were given chronic intermittent mild stress (CIMS) exposure (3 cycles, in which each cycle consists of once daily mild stress for 5 days/week for 2 weeks, followed by 2 weeks of no stress) vs. non-stress controls, combined with ongoing access to a palatable diet (PD; choice of chow, high-fat diet, 30% sucrose drink, and water) vs. control diet (chow and water). As expected, access to PD increased caloric intake, body weight gain, and adiposity, and impaired glucose tolerance. CIMS decreased body weight gain only during the first cycle of stress and did not affect body weight gain thereafter, regardless of diet. Moreover, CIMS did not alter total food intake, adiposity or glucose tolerance regardless of diet. Lastly, CIMS transiently increased high-fat diet preference in PD-fed rats during the first stress cycle. Collectively, these results suggest that CIMS has relatively modest metabolic effects that occur primarily during initial stress exposure. These results support the hypothesis that the metabolic consequences of chronic stress vary with stress intensity and/or frequency.

Weight loss by calorie restriction versus bariatric surgery differentially regulates the hypothalamo-pituitary-adrenocortical axis in male rats.

Behavioral modifications for the treatment of obesity, including caloric restriction, have notoriously low long-term success rates relative to bariatric weight-loss surgery. The reasons for the difference in sustained weight loss are not clear. One possibility is that caloric restriction alone activates the stress-responsive hypothalamo-pituitary-adrenocortical (HPA) axis, undermining the long-term maintenance of weight loss, and that this is abrogated after bariatric surgery. Accordingly, we compared the HPA response to weight loss in five groups of male rats: (1) high-fat diet-induced obese (DIO) rats treated with Roux-en-Y gastric bypass surgery (RYGB, n = 7), (2) DIO rats treated with vertical sleeve gastrectomy (VSG, n = 11), (3) DIO rats given sham surgery and subsequently restricted to the food intake of the VSG/RYGB groups (Pair-fed, n = 11), (4) ad libitum-fed DIO rats given sham surgery (Obese, n = 11) and (5) ad libitum chow-fed rats given sham surgery (Lean, n = 12). Compared with Lean controls, food-restricted rats exhibited elevated morning (nadir) non-stress plasma corticosterone concentration and increased hypothalamic corticotropin-releasing hormone and vasopressin mRNA expression, indicative of basal HPA activation. This was largely prevented when weight loss was achieved by bariatric surgery. DIO increased HPA activation by acute (novel environment) stress and this was diminished by bariatric surgery-, but not pair-feeding-, induced weight loss. These results indicate that the HPA axis is differentially affected by weight loss from caloric restriction versus bariatric surgery, and this may contribute to the differing long-term effectiveness of these two weight-loss approaches.

Loss of melanocortin-4 receptor function attenuates HPA responses to psychological stress.

The melanocortin 4 receptor (MC4R), well-known for its role in the regulation of energy balance, is widely expressed in stress-regulatory brain regions, including the paraventricular nucleus of the hypothalamus (PVH) and the medial amygdala (MeA). In agreement with this, MC4R has been implicated in hypothalamic-pituitary-adrenocortical axis (HPA) regulation. The present work investigated the role of chronic Mc4r function to modulate basal HPA axis tone and to facilitate acute HPA responses to psychological stress, using a novel rat model with Mc4r loss-of-function. In this study, adult male rats were placed into 3 groups (n=15/group) according to genotype [wild-type (WT); heterozygous mutant (HET); and homozygous mutant (HOM)]. Basal (pre-stress) plasma adrenocorticotropic hormone (ACTH) and corticosterone were measured in the AM and PM, and the HPA axis response to restraint was assessed in the AM. Rats were perfused at 2h after restraint to assess the effect of loss of MC4R on stress-induced c-Fos immunolabeling in stress-regulatory brain regions. We find that basal (non-stress) AM and PM plasma ACTH and corticosterone showed a normal diurnal rhythm that was not altered according to genotype. Consistent with this, adrenal and thymus weights were unaffected by genotype. However, the plasma ACTH and corticosterone responses to restraint were significantly reduced by loss of MC4R function. Likewise, stress-induced c-Fos immunolabeling in both PVH and MeA was significantly reduced by loss of Mc4r function. These results support the hypothesis that endogenous MC4R signaling contributes to the HPA axis response to stress. Because MC4R plays a critical role in the regulation of energy balance, the present work suggests that it may also serve as an important communication link between brain metabolic and stress systems.

Morphological and behavioral impact of AAV2/5-mediated overexpression of human wildtype alpha-synuclein in the rat nigrostriatal system.

The discovery of the involvement of alpha-synuclein (α-syn) in Parkinson's disease (PD) pathogenesis has resulted in the development and use of viral vector-mediated α-syn overexpression rodent models. The goal of these series of experiments was to characterize the neurodegeneration and functional deficits resulting from injection of recombinant adeno-associated virus (rAAV) serotype 2/5-expressing human wildtype α-syn in the rat substantia nigra (SN). Rats were unilaterally injected into two sites in the SN with either rAAV2/5-expressing green fluorescent protein (GFP, 1.2 x 10(13)) or varying titers (2.2 x 10(12), 1.0 x 10(13), 5.9 x 10(13), or 1.0 x 10(14)) of rAAV2/5-α-syn. Cohorts of rats were euthanized 4, 8, or 12 weeks following vector injection. The severity of tyrosine hydroxylase immunoreactive (THir) neuron death in the SN pars compacta (SNpc) was dependent on vector titer. An identical magnitude of nigrostriatal degeneration (60-70% SNpc THir neuron degeneration and 40-50% loss of striatal TH expression) was observed four weeks following 1.0 x 10(14) titer rAAV2/5-α-syn injection and 8 weeks following 1.0 x 10(13) titer rAAV2/5-α-syn injection. THir neuron degeneration was relatively uniform throughout the rostral-caudal axis of the SNpc. Despite equivalent nigrostriatal degeneration between the 1.0 x 10(13) and 1.0 x 10(14) rAAV2/5-α-syn groups, functional impairment in the cylinder test and the adjusting steps task was only observed in rats with the longer 8 week duration of α-syn expression. Motor impairment in the cylinder task was highly correlated to striatal TH loss. Further, 8 weeks following 5.9 x 10(13) rAAV2/5-α-syn injection deficits in ultrasonic vocalizations were observed. In conclusion, our rAAV2/5-α-syn overexpression model demonstrates robust nigrostriatal α-syn overexpression, induces significant nigrostriatal degeneration that is both vector and duration dependent and under specific parameters can result in motor impairment that directly relates to the level of striatal TH denervation.