N6-methyladenine is an epigenetic marker of mammalian early life stress.

Recent evidence described 6-methyladenine (6 mA) as a novel epigenetic regulator in a variety of multicellular species, including rodents; however, its capacity to influence gene expression in the mammalian brain remains unknown. We examined if 6 mA is present and regulated by early life stress associated with predator odor exposure (POE) within the developing rat amygdala. Our results provide evidence that 6 mA is present in the mammalian brain, is altered within the Htr2a gene promoter by early life stress and biological sex, and increased 6 mA is associated with gene repression. These data suggest that methylation of adenosine within mammalian DNA may be used as an additional epigenetic biomarker for investigating the development of stress-induced neuropathology.

Antipsychotic-like actions of the satiety peptide, amylin, in ventral striatal regions marked by overlapping calcitonin receptor and RAMP-1 gene expression.

Amylin is a calcitonin-related peptide co-secreted with insulin, which produces satiety through brainstem-localized receptors; however, its effects in forebrain are poorly understood. The nucleus accumbens shell (AcbSh) exhibits among the densest concentrations of high-affinity amylin binding; nevertheless, these receptors have not been explored beyond one study showing dopamine antagonist-like effects of intra-Acb amylin on feeding and associated behavior (Baldo and Kelley, 2001). Here, we investigated whether intra-Acb amylin signaling modulates prepulse inhibition (PPI), a measure of sensorimotor gating deficient in several illnesses including schizophrenia. First, in situ hybridization revealed marked anatomical gradients for both receptor activity-modifying protein-1 (RAMP-1) and calcitonin receptor gene (CT-R) expression in striatum [coexpression of these genes yields a high-affinity amylin-1 receptor (AMY1-R)], with highest overlap in the medial AcbSh. Intra-AcbSh amylin infusions in rats (0, 30, and 100 ng) reversed amphetamine (AMPH)-induced PPI disruption without affecting baseline startle; dorsal striatal amylin infusions had no effect. Coinfusion of AC187 (20 µg), an antagonist for AMY1-R, blocked the ability of amylin to normalize AMPH-induced PPI disruption, showing the specificity of AcbSh amylin effects to the AMY1-R. Intra-AcbSh AC187 on its own disrupted PPI in a haloperidol-reversible manner (0.1 mg/kg). Thus, AMY1-R may be a potential target for the development of putative antipsychotics or adjunct treatments that oppose metabolic side effects of current medications. Moreover, AMY1-Rs may represent a novel way to modulate activity preferentially in ventral versus dorsal striatum.

Principles of motivation revealed by the diverse functions of neuropharmacological and neuroanatomical substrates underlying feeding behavior.

Circuits that participate in specific subcomponents of feeding (e.g., gustatory perception, peripheral feedback relevant to satiety and energy balance, reward coding, etc.) are found at all levels of the neural axis. Further complexity is conferred by the wide variety of feeding-modulatory neurotransmitters and neuropeptides that act within these circuits. An ongoing challenge has been to refine the understanding of the functional specificity of these neurotransmitters and circuits, and there have been exciting advances in recent years. We focus here on foundational work of Dr. Ann Kelley that identified distinguishable actions of striatal opioid peptide modulation and dopamine transmission in subcomponents of reward processing. We also discuss her work in overlaying these neuropharmacological effects upon anatomical pathways that link the telencephalon (cortex and basal ganglia) with feeding-control circuits in the hypothalamus. Using these seminal contributions as a starting point, we will discuss new findings that expand our understanding of (1) the specific, differentiable motivational processes that are governed by central dopamine and opioid transmission, (2) the manner in which other striatal neuromodulators, specifically acetylcholine, endocannabinoids and adenosine, modulate these motivational processes (including via interactions with opioid systems), and (3) the organization of the cortical-subcortical network that subserves opioid-driven feeding. The findings discussed here strengthen the view that incentive-motivational properties of food are coded by substrates and neural circuits that are distinguishable from those that mediate the acute hedonic experience of food reward. Striatal opioid transmission modulates reward processing by engaging frontotemporal circuits, possibly via a hypothalamic-thalamic axis, that ultimately impinges upon hypothalamic modules dedicated to autonomic function and motor pattern control. We will conclude by discussing implications for understanding disorders of "non-homeostatic" feeding.

Fos expression following regimens of predator stress versus footshock that differentially affect prepulse inhibition in rats.

Stress is suggested to exacerbate symptoms and contribute to relapse in patients with schizophrenia and several other psychiatric disorders. A prominent feature of many of these illnesses is an impaired ability to filter information through sensorimotor gating processes. Prepulse inhibition (PPI) is a functional measure of sensorimotor gating, and known to be deficient in schizophrenia and sometimes in post-traumatic stress disorder (PTSD), both of which are also sensitive to stress-induced symptom deterioration. We previously found that a psychological stressor (exposure to a ferret without physical contact), but not footshock, disrupted PPI in rats, suggesting that intense psychological stress/trauma may uniquely model stress-induced sensorimotor gating abnormalities. In the present experiment, we sought to recreate the conditions where we found this behavioral difference, and to explore possible underlying neural substrates. Rats were exposed acutely to ferret stress, footshock, or no stress (control). 90 min later, tissue was obtained for Fos immunohistochemistry to assess neuronal activation. Several brain regions (prelimbic, infralimbic, and cingulate cortices, the paraventricular hypothalamic nucleus, the paraventricular thalamic nucleus, and the lateral periaqueductal gray) were equally activated following exposure to either stressor. Interestingly, the medial amygdala and dorsomedial periaqueductal gray had nearly twice as much Fos activation in the ferret-exposed rats as in the footshock-exposed rats, suggesting that higher activation within these structures may contribute to the unique behavioral effects induced by predator stress. These results may have implications for understanding the neural substrates that could participate in sensorimotor gating abnormalities seen in several psychiatric disorders after psychogenic stress.

Predator threat induces behavioral inhibition, pituitary-adrenal activation and changes in amygdala CRF-binding protein gene expression.

Behavioral inhibition (BI) is an adaptive defensive response to threat; however, extreme BI is associated with anxiety-related psychopathology. When rats are exposed to a natural predator they display stress- and anxiety-related behavioral alterations and physiological activation. To develop a preclinical rodent model to study mechanisms underlying human BI and anxiety, we examined the extent to which ferret exposure elicits anxiety-related BI and HPA and amygdala activation of the CRF system. In the first experiment, BI and other behaviors were assessed in the presence or absence of a ferret. In the second experiment, ferret-induced corticosterone release and changes in brain c-fos expression were assessed. In the final experiment, gene chip and quantitative real time-PCR analyses were performed on amygdala tissue from control and ferret-exposed rats. Ferret exposure increased BI and submissive posturing, as well as plasma corticosterone and the number of Fos-positive cells in several brain regions including the amygdala. Gene expression analysis revealed increased amygdalar mRNA for CRF-binding protein, but not the CRF1 receptor, CRF2 receptor or CRF. In rodents, ferret exposure can be used to elicit anxiety-related BI, which is associated with HPA and amygdala activation. Since the amygdala and the CRF system have been implicated in adaptive and maladaptive anxiety responses in humans, these data support use of our rodent model to further investigate mechanisms underlying anxiety-related psychopathology in humans.

Reduction of stress-induced behavior by antagonism of corticotropin-releasing hormone 2 (CRH2) receptors in lateral septum or CRH1 receptors in amygdala.

Although corticotropin-releasing hormone (CRH), a regulator of stress responses, acts through two receptors (CRH1 and CRH2), the role of CRH2 in stress responses remains unclear. Knock-out mice without the CRH2 gene exhibit increased stress-like behaviors. This profile could result either directly from the absence of CRH2 receptors or indirectly from developmental adaptations. In the present study, CRH2 receptors were acutely blocked by alpha-helical CRH (alpha(h)CRH, CRH1/CRH2 antagonist; 0, 30, 100, and 300 ng) infusion into the lateral septum (LS), which abundantly expresses CRH2 but not CRH1 receptors. Freezing, locomotor activity, and analgesia were tested after infusion. Intra-LS alpha(h)CRH blocked shock-induced freezing without affecting activity or pain responses; infusions into lateral ventricle or nucleus of the diagonal band had no effects. The same behavioral profile was obtained with d-Phe-CRH((12-41)) (100 ng), another CRH1/CRH2 antagonist. A selective CRH1 antagonist (NBI27914), in doses that reduced freezing on intra-amygdala (central nucleus) infusion (0, 0.2, and 1.0 microg), did not affect freezing when infused into the LS. Ex vivo autoradiography revealed that binding of [125I]sauvagine, a mixed CRH1/CRH2 agonist, was prevented in the LS by previous intra-LS infusion of alpha(h)CRH but not NBI27914. In vitro studies demonstrated that [125I]sauvagine binding in the LS could be inhibited by a CRH1/CRH2 antagonist but not by the selective CRH1 receptor antagonist, confirming that in the LS, alpha(h)CRH antagonized exclusively CRH2 receptors. Acute antagonism of CRH2 receptors in the LS thus produces a behaviorally, anatomically, and pharmacologically specific reduction in stress-induced behavior, in contrast to results of recent knock-out studies, which induced congenital and permanent CRH2 removal. CRH2 receptors may thus represent a potential target for the development of novel CRH system anxiolytics.