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It is widely accepted that leptin acts on first-order neurons in the arcuate nucleus (ARC) with information then relayed to other hypothalamic centers. However, the extent to which leptin mediates its central actions solely, or even primarily, via this route is unclear. We used a model of hypothalamo-pituitary disconnection (HPD) to determine whether leptin action on appetite-regulating systems requires the ARC. This surgical preparation eliminates the ARC. We measured effects of iv leptin to activate hypothalamic neurons (Fos labeling). In ARC-intact animals, leptin increased the percentage of Fos-positive melanocortin neurons and reduced percentages of Fos-positive neuropeptide Y neurons compared with saline-treated animals. HPD itself increased Fos labeling in the lateral hypothalamic area (LHA). Leptin influenced Fos labeling in the dorsomedial nucleus (DMH), ventromedial nucleus, and paraventricular nucleus (PVN) in HPD and normal animals, with effects on particular cell types varying. In the LHA and DMH, leptin decreased orexin cell activation in HPD and ARC-intact sheep. HPD abolished leptin-induced expression of Fos in melanin-concentrating hormone cells in the LHA and in CRH cells in the PVN. In contrast, HPD accentuated activation in oxytocin neurons. Our data from sheep with lesions encompassing the ARC do not suggest a primacy of action of leptin in this nucleus. We demonstrate that first order to second order signaling may not represent the predominant means by which leptin acts in the brain to generate integrated responses. We provide evidence that leptin exerts direct action on cells of the DMH, ventromedial nucleus, and PVN.

Galanin (GAL) stimulates food intake in normal rats when it is injected in different hypothalamic areas involved in feeding such as the paraventricular and ventromedial nuclei and the lateral hypothalamus. At adulthood, the hyperphagic obese Zucker rat is characterized by a general dysregulation of some important neuropeptides involved in the regulation of food intake including GAL. The aim of this study was to measure GAL in different microdissected brain areas in 2- and 4-week-old lean (FA/-) and obese (fa/fa) male Zucker rats in order to know if GAL actively participates in triggering abnormal feeding behavior in obese rats. There was a significant increase (40%-220%) in GAL concentration with age in the arcuate and dorsomedial nuclei and in the above areas except for the lateral hypothalamus. Genotype differences were observed in the arcuate and paraventricular nuclei only. GAL levels were globally lower in obese than in lean rats (-15% to -25%) and the difference was significant at 2 weeks of age in the paraventricular nucleus and at 4 weeks of age in the arcuate nucleus. In agreement with human observations, these data suggest that GAL is not an early player in the development of overeating.

In the present study, we investigated the role of prolactin (PRL) in the suckling-induced increase in hypothalamic neuropeptide Y (NPY) gene expression in the dorsomedial nucleus of the hypothalamus (DMH) and the caudal portion of the arcuate nucleus of the hypothalamus (ARH-C). Lactating rats were deprived of their eight-pup litters on d 9 postpartum. After 48 h, the animals were randomly divided into two groups: nonsuckled controls and eight pups suckling for 24 h. In addition, some of the suckled animals received two injections of bromocriptine (0.5 mg/rat per injection) to inhibit suckling-induced PRL secretion. Some bromocriptine-treated rats also received ovine PRL (1 mg/rat per injection). In situ hybridization was performed to measure NPY mRNA levels. Suckling for 24 h induced a significant increase in NPY mRNA levels in the DMH and ARH-C. Bromocriptine treatment greatly attenuated the increase of NPY mRNA in the DMH but not in the ARH. Injections of ovine PRL in bromocriptine-treated rats greatly restored DMH NPY mRNA levels but had no additional effects on the ARH NPY expression. Double-label in situ hybridization for NPY and PRL receptor (PRL-R) in the lactating rat brains showed that NPY-positive neurons in the DMH also express PRL-R mRNA. On the contrary, few ARH NPY neurons expressed PRL-R. These data suggest that PRL could act directly on DMH NPY neurons to modulate NPY gene expression during lactation. Thus, the results from the present study demonstrate that NPY neurons in the DMH and ARH are differentially regulated by PRL during lactation.

Gamma-aminobutyric acid (GABA) interacts with hypothalamic neuronal pathways regulating feeding behaviour. GABA has been reported to stimulate feeding via both ionotropic GABA(A) and metabotropic GABA(B) receptors. The functional form of the GABA(B) receptor is a heterodimer consisting of GABA(B) receptor-1 (GABA(B)R1) and GABA(B) receptor-2 (GABA(B)R2) proteins. Within the heterodimer, the GABA-binding site is localized to GABA(B)R1. In the present study, we used an antiserum to the GABA(B)R1 protein in order to investigate the cellular localization of GABA(B)R1-immunoreactive neurones in discrete hypothalamic regions implicated in the control of body weight. The colocalization of GABA(B)R1 immunoreactivity with different chemical messengers that regulate food intake was analysed. GABA(B)R1-immunoreactive cell bodies were found in the periventricular, paraventricular (PVN), supraoptic, arcuate, ventromedial hypothalamic, dorsomedial hypothalamic, tuberomammillary nuclei and lateral hypothalamic area (LHA). Direct double-labelling showed that glutamic acid decarboxylase (GAD)-positive terminals were in close contact with GABA(B)R1-containing cell bodies located in all these regions. In the ventromedial part of the arcuate nucleus, GABA(B)R1-immunoreactive cell bodies were found to contain neuropeptide Y, agouti-related peptide (AGRP) and GAD. In the ventrolateral part of the arcuate nucleus, GABA(B)R1-immunoreactive cell bodies were shown to contain pro-opiomelanocortin and cocaine- and amphetamine-regulated transcript. In the LHA, GABA(B)R1 immunoreactivity was present in both melanin-concentrating hormone- and orexin-containing cell populations. In the tuberomammillary nucleus, GABA(B)R1-immunoreactive cell bodies expressed histidine decarboxylase, a marker for histamine-containing neurones. In addition, GAD and AGRP were found to be colocalized in some nerve terminals surrounding GABA(B)R1-immunoreactive cell bodies in the parvocellular part of the PVN. The results may provide a morphological basis for the understanding of how GABA regulates the hypothalamic control of food intake and body weight via GABA(B) receptors.

Hypothalamic neuropeptide Y (NPY) and corticotropin-releasing hormone (CRH) influence feeding and levels of plasma glucose, insulin, free fatty acids, and triglycerides. Treatment of genetically obese, ob/ob mice, with dopamine receptor D(1)/D(2) agonists normalizes hyperphagia, body weight gain, hyperglycemia, and hyperlipidemia. We therefore examined whether levels of NPY and CRH immunoreactivity in discrete hypothalamic nuclei are altered in ob/ob mice, and whether dopaminergic treatment reverses this alteration. Female ob/ob mice were treated daily at 1 h after light onset with the D(1)/D(2) agonists, SKF-38393 (20 mg/kg) and bromocriptine (15 mg/kg), respectively or vehicle for 2 weeks. Such treatment, while normalizing body weight gain and hyperglycemia, also significantly reduced elevated NPY immunoreactivity in the suprachiasmatic (by 39%), intergeniculate (by 43%), paraventricular (PVN; by 31%), and arcuate (by 41%) nuclei in obese mice to levels observed in lean mice. This treatment also caused a 45-50% decline in levels of CRH in the PVN and dorsomedial hypothalamus compared to obese controls to levels observed in lean mice. Taken together, these findings suggest that dopaminergic D(1)/D(2) receptor coactivation may improve hyperphagia, hyperglycemia, and obesity in the ob/ob mouse, in part, by normalizing elevated levels of both NPY and CRH., (Copyright 2000 S. Karger AG, Basel.)

A newly identified hypothalamic peptide whose specific receptors are present in the anterior pituitary gland is a selective and potent stimulator of prolactin secretion and is therefore termed prolactin-releasing peptide (PRP). We investigated the distribution of PRP-containing neurons in the hypothalamus of female rats by immunocytochemical techniques. Immunocytochemistry using a specific antibody raised to PRP revealed that PRP-immunoreactive perikarya were located in the posteroventral part of the dorsomedial nucleus of the hypothalamus. PRP-immunoreactive nerve terminals were present in high concentrations in a region ventral to the bed nucleus of the stria terminalis, but scarcely observed in the external layer of the median eminence in which well known hypothalamic hormones such as growth hormone-releasing hormone and somatostatin were abundantly detected. This specific distribution in the hypothalamus suggests a novel route of the hypophysiotropic action of PRP.

The wild white-footed mouse, Peromyscus leucopus, is commonly used for photoperiod studies utilizing physiological, behavioral, and other biological measures indicative of hypothalamic functions. Indoleamines, like melatonin and serotonin, are implicated in regulating these hypothalamic functions. Although neurochemical analyses of hypothalamic serotonin and its receptors have been reported for this species, the relevant neuroanatomy of the serotonin system within mouse hypothalamus has not been studied. A sensitive immunohistochemical method was used to detect serotonin within axons of coronal sections of formaldehyde fixed forebrain from P. leucopus. Large, medium and small diameter serotonin axons were evaluated in most regions, or nuclei, of the hypothalamus rostral to the mammillary region. A fourth type of serotonin axon was observed to have morphology characteristic of terminal arbors. The density of serotonin axons ranged from no staining to very high density similar to other species for which reports exist, i.e., rat, cat, and monkey. The ventromedial hypothalamic nucleus had distinctively lesser density of serotonin axons in this mouse than other species. Evidence of terminal arborization in hypothalamic nuclei and regions was evident. Neuroendocrine, autonomic, and behavioral functions of the hypothalamus are suggested to be regulated by input from serotonin terminals in this wild species of mouse, in correlation with receptor localization as reported by others., (Copyright 1998 Elsevier Science B.V.)

Regional hypothalamic neuropeptide Y (NPY) concentrations were compared between cp/cp JCR:LA corpulent rats, which were grossly obese, hyperphagic, and hyperinsulinemic, and lean (+/+) controls. In freely fed cp/cp rats, NPY levels in the arcuate nucleus (ARC) were 31% higher than in lean rats (p less than 0.001). In lean rats, chronic food restriction significantly raised NPY levels by 22% in the ARC (p less than 0.05) and by 44% in the dorsomedial nucleus (DMH; p less than 0.05). By contrast, food-restricted cp/cp rats showed no change in the ARC, but NPY levels rose in the DMH (by 36%; p less than 0.05) and ventromedial nucleus (31%; p less than 0.05). Increased NPY levels in the ARC, the major site of hypothalamic NPY synthesis, suggests increased NPYergic activity in cp/cp rats; given the central actions of NPY, this could contribute to hyperphagia, obesity, and hyperinsulinemia in this syndrome. Abnormal NPY responses to food deprivation further suggest dysregulation of NPY in cp/cp rats.

Neuropeptide Y (NPY) concentrations were measured by radioimmunoassay in eight microdissected hypothalamic regions of obese (fa/fa) and lean (Fa/?) Zucker rats. Freely fed obese rats showed significant (40-100%) increases in NPY concentrations in several regions, notably the paraventricular, ventromedial, and dorsomedial nuclei and the arcuate nucleus/median eminence, compared with lean rats. Hypothalamic NPY concentrations were not affected in either obese or lean rats by food restriction, which caused 25% weight loss over 3 wk. Refeeding to initial weight significantly increased NPY levels in the ventromedial and dorsomedial nuclei in lean rats but did not significantly alter NPY concentrations in any hypothalamic region in obese rats. These observations indicate fundamental differences in the regulation of hypothalamic NPY between obese and lean Zucker rats. NPY injected into the paraventricular nucleus and other regions causes hyperphagia, obesity, and increased secretion of insulin, glucagon, ACTH, and corticosterone. These behavioral and neuroendocrine abnormalities all occur in the obese Zucker syndrome and may be due to increased NPY-ergic activity in the hypothalamus.