Efferent and afferent connections of the ventromedial hypothalamic nucleus determined by neural tracer analysis: implications for lordosis regulation in female rats.

Neural connections of the ventromedial hypothalamic nucleus (VMN) to and from forebrain and midbrain structures, which are involved in the neuroendocrine regulation of reproduction, were investigated. A retrograde (fluoro-gold [FG]) or an anterograde neural tracer (phaseolus vulgaris-leucoagglutinin [PHA-L]) was injected into the left side of the VMN in ovariectomized rats. Six days after injection with FG or 11 days after injection with PHA-L, brains were fixed and sectioned. After immunohistochemistry, digital images of FG-labeled neural cell bodies (FG-cells) or PHA-L-labeled fibers (PHA-L-fibers) were analyzed. Injection sites of FG and PHA-L were mainly in the ventrolateral VMN. Considerable numbers of FG-cells and PHA-L-fibers were present in the left side of the medial amygdala, ventral lateral septum, preoptic area, bed nucleus of stria terminalis, dorsomedial hypothalamic nucleus, arcuate nucleus, periventricular nucleus of thalamus, and midbrain central gray. The lateral dorsal raphe nuclei contained many PHA-L-fibers but few FG-cells. By contrast, both sides of the median raphe nucleus contained many FG-cells but few PHA-L-fibers. Reciprocal direct neural connection between the right and left side of the VMN were observed. The present results provide an anatomical basis for functional relationships between the VMN and these nuclei.

Identification of urocortin 3 afferent projection to the ventromedial nucleus of the hypothalamus in rat brain.

Urocortin 3 (Ucn 3) is a corticotrophin-releasing factor related neuropeptide highly expressed in the brain. Ucn 3 nerve fibers heavily innervate the hypothalamic ventromedial nucleus (VMH), and Ucn 3 injection into the VMH suppresses feeding. Currently, the origin of the Ucn 3 afferent input into the VMH is unknown. In the present study, anatomical tracing shows that the major Ucn 3 afferent input to the VMH resides in the anterior parvicellular part of the paraventricular nucleus of the hypothalamus (PVHap) and the adjacent posterior part of the bed nucleus of stria terminalis (pBNST). VMH also receives moderate Ucn 3 input from the medial amygdala. Ucn 3 neurons located immediately caudal to the PVHap/pBNST in the rostral perifornical hypothalamic area (rPFH) provide only minimal input. The paucity of rPFH-VMH Ucn 3 projection is consistent with the finding that only Ucn 3 neurons in the rPFH co-expressed enkephalin (Enk), and Ucn 3/Enk double-labeled nerve fibers and terminals were observed predominately in the lateral septum (LS), whereas only a few double-labeled fibers were found in other brain areas including the VMH. Furthermore, retrograde tracing demonstrates that Ucn 3 neurons in the rPFH project to the LS. In conclusion, the present study determines that the major Ucn 3 afferent into the VMH originates from the PVHap/pBNST. Moreover, anatomical heterogeneity is observed in the hypothalamic Ucn 3 neuron population as the rostral part (PVHap/pBNST) of the population projects to the VMH and the caudal part (rPFH) co-localizes with Enk and provides major afferent input to the LS.

Functional identification of an aggression locus in the mouse hypothalamus.

Electrical stimulation of certain hypothalamic regions in cats and rodents can elicit attack behaviour, but the exact location of relevant cells within these regions, their requirement for naturally occurring aggression and their relationship to mating circuits have not been clear. Genetic methods for neural circuit manipulation in mice provide a potentially powerful approach to this problem, but brain-stimulation-evoked aggression has never been demonstrated in this species. Here we show that optogenetic, but not electrical, stimulation of neurons in the ventromedial hypothalamus, ventrolateral subdivision (VMHvl) causes male mice to attack both females and inanimate objects, as well as males. Pharmacogenetic silencing of VMHvl reversibly inhibits inter-male aggression. Immediate early gene analysis and single unit recordings from VMHvl during social interactions reveal overlapping but distinct neuronal subpopulations involved in fighting and mating. Neurons activated during attack are inhibited during mating, suggesting a potential neural substrate for competition between these opponent social behaviours.

Noradrenergic nuclei that receive sensory input during mating and project to the ventromedial hypothalamus play a role in mating-induced pseudopregnancy in the female rat.

In female rats, vaginal-cervical stimulation (VCS) received during mating induces bicircadian prolactin surges that are required for the maintenance of pregnancy or pseudopregnancy (PSP). The neural circuits that transmit VCS inputs to the brain have not been fully described, although mating stimulation is known to activate medullary noradrenergic cell groups that project to the forebrain. In response to VCS, these neurones release noradrenaline within the ventrolateral division of the ventromedial hypothalamus (VMHvl) and the posterodorsal medial amygdala (MePD), two forebrain sites that are implicated in the initiation of PSP. Noradrenaline receptor activation within the VMHvl is both necessary and sufficient for PSP induction, suggesting that noradrenaline acting within the VMHvl is particularly important in mediating the effects of VCS towards the establishment of PSP. We therefore investigated whether or not endogenous, VCS-induced noradrenaline release within the VMHvl is involved in PSP induction in the rat. Before the receipt of sufficient mating stimulation to induce PSP, a retrograde neurotoxin, dopamine-ß-hydroxylase-saporin (DBH-SAP), was infused bilaterally into the either the VMHvl or the MePD to selectively destroy afferent noradrenergic nuclei in the brainstem. DBH-SAP infusions into the VMHvl lesioned mating-responsive noradrenergic neurones in A1 and A2 medullary nuclei and reduced the incidence of PSP by 50%. Infusions of DBH-SAP into the MePD had no effect on the subsequent induction of PSP. These results suggest that VCS is conveyed to mating-responsive forebrain areas by brainstem noradrenergic neurones, and that the activity of noradrenergic cells projecting to the VMHvl is involved in the induction of PSP.

Relationship of arousal to circadian anticipatory behavior: ventromedial hypothalamus: one node in a hunger-arousal network.

The mechanisms by which animals adapt to an ever-changing environment have long fascinated scientists. Different forces, conveying information regarding various aspects of the internal and external environment, interact with each other to modulate behavioral arousal. These forces can act in concert or, at times, in opposite directions. These signals eventually converge and are integrated to influence a common arousal pathway which, depending on all the information received from the environment, supports the activation of the most appropriate behavioral response. In this review we propose that the ventromedial hypothalamic nucleus (VMN) is part of the circuitry that controls food anticipation. It is the first nucleus activated when there is a change in the time of food availability, silencing of VMN ghrelin receptors decreases food-anticipatory activity (FAA) and, although lesions of the VMN do not abolish FAA, parts of the response are often altered. In proposing this model it is not our intention to exclude parallel, redundant and possibly interacting pathways that may ultimately communicate with, or work in concert with, the proposed network, but rather to describe the neuroanatomical requirements for this circuit and to illustrate how the VMN is strategically placed and connected to mediate this complex behavioral adaptation.

The effects of partial and complete masculinization on the sexual differentiation of nuclei that control lordotic behavior in the male rat.

Male rats, under certain experimental conditions, may show lordosis, the typical expression of female sexual receptivity. This work studies the sexual morphological pattern of facilitatory and inhibitory structures that control lordosis. Three groups of males were neonatally subjected to a gradient of androgen exposure (castrated plus injected oil (GxM+oil); castrated plus androstenedione treated (GxM+AND); and sham operated [CM]); a group of control females (CF) was also added. Lordotic response after these different hormonal and neonatal surgical treatments, as well as the volume or number of neurons in facilitatory (ventromedial nucleus of the hypothalamus [VMN]) and inhibitory (the intermediate region of the lateral septum [LSi] and accessory olfactory bulb [AOB]) nuclei involved in lordosis was studied in adults. The inhibition of lordosis in the males seems to be associated to the neonatal presence of testosterone and the consequent masculinization of the VMN, VMNvl, LSi and AOB. It is suggested that one of the functions of the sex differences consistently seen in these structures might be to inhibit the lordosis response in the male.

Sex and seasonal differences in morphology of limbic forebrain nuclei in the green anole lizard.

Sex and seasonal differences in the brain occur in many species and are often related to behavioral expression. For example, morphology of limbic regions involved in male sex behavior are larger in males than in females, and sometimes are larger in the breeding than non-breeding season. Morphology can often be altered in adulthood by manipulating levels of steroid hormones. In untreated green anole lizards, previous work indicated that neuron soma size and density did not differ between the sexes in the preoptic area (POA) or ventromedial nucleus of the amygdala (AMY), two brain regions involved in the control of male reproductive behaviors [O'Bryant, E.L., Wade, J., 2002. Seasonal and sexual dimorphisms in the green anole forebrain. Horm. Behav. 41, 384-395.]. However, soma size was larger in both areas in breeding than non-breeding animals. The current study examined sex and seasonal differences in estimated brain region volume and total neuron number in the POA, AMY, and the ventromedial hypothalamus (VMH), a region typically involved in female reproductive behaviors. The volume of the POA was larger in males, and the POA and VMH of breeding animals were larger than those of non-breeding individuals. Differences in cell number did not exist in either of these two regions. In contrast, neuron counts in the AMY were greater in non-breeding than breeding animals, but the volume did not differ between the seasons. These data suggest that the structure of limbic brain regions is dynamic in adulthood and that parallels between morphology and the expression of masculine behavior exist for the POA, whereas other relationships are more complicated.

Ventromedial hypothalamic NPY Y2 receptor in the maintenance of body weight in diet-induced obesity in mice.

This study examined changes in neuropeptide Y (NPY) Y2 receptor binding in the brains of C57BL/6 mice in response to different levels of high-fat diets via three dietary intervention methods: high-fat diet, switching from high- to low-fat diet and finally, energy restricted high-fat diet. Forty-five C57Bl/6 male mice were fed a high-fat diet for 8 weeks and then classified as diet-induced obese (DIO) or diet-resistant (DR) mice according to the highest and lowest body weight gainers, respectively. The DIO and DR mice were then randomly divided into three groups each and either continued on their high-fat diet ad libitum (DIO-H and DR-H), changed to a low-fat diet (DIO-L and DR-L) or pair-fed via energy restricted high-fat diet (DIO-P and DR-P) for a further 6 weeks. During the course of this study, body weight, energy intake and plasma peptide YY (PYY) were measured. The study revealed that the replacement of a high-fat diet with a low-fat diet was associated with a significant lowering of ventromedial hypothalamic (VMH) Y2 receptor binding in both the DIO-L and DR-L mice (-37%, -36%), and also a lowered plasma PYY level in the DIO-L mice (-25%). Despite a continued consumption of the high-fat diet, energy restricted pair feeding caused a lower VMH Y2 receptor binding in the obese mice (DIO-P) following weight loss compared to the DR-P mice (-14%). In conclusion, this study showed that changing diets from high- to low-fat can significantly lower the VMH Y2 receptor binding irrespective to the obesity phenotype. Energy restriction, even while on high-fat feeding, can cause a lower VMH Y2 receptor binding compared to DR mice even after body weight loss to similar levels. This suggests either a possible intrinsic nature of the DIO mice or a body weight set-point re-establishment to drive body weight regain.

The effects of early isolation on sexual behavior and c-fos expression in naïve male Long-Evans rats.

Previous findings have demonstrated that the maternal environment is important for the development of male sexual behavior. The present study examined the effects of complete early life isolation and replacement 'stroking' stimulation on male sexual behavior and neural activation as seen by Fos immunoreactivity (Fos-IR). Animals were either artificially reared (AR) with minimal (AR-MIN) or maximal (AR-MAX) body simulation, or maternally reared (MR). In adulthood, animals were either given an exposure to an estrous female (EXP) or left undisturbed (NoEXP). No significant effects of early development were found in sexual behavior; however differences in activation in response to this exposure were observed. AR-MIN animals showed lower Fos-IR in the medial preoptic area and the ventromedial hypothalamus compared to MR animals. AR-MAX animals were not significantly different from either condition. These findings demonstrate that although there are no differences in the quality of the first copulatory exposure between AR and MR animals, the brain's response to this exposure differs in sites within the brain that subserve sexual behavior.

Caudal hindbrain lactate infusion alters glucokinase, SUR1, and neuronal substrate fuel transporter gene expression in the dorsal vagal complex, lateral hypothalamic area, and ventromedial nucleus hypothalamus of hypoglycemic male rats.

While in vitro studies show that the oxidizable energy substrate, lactate, is a preferred fuel for CNS neurons during states of energy crisis, and that lactate may regulate neuronal glucose uptake under those conditions, its role in neuronal function in vivo remains controversial. Glucose-excited neurons in hindbrain dorsal vagal complex (DVC) monitor both glucose and lactate, and express both the glucose sensor, glucokinase (GK), and the SUR1 subunit of the plasma membrane energy transducer, K(ATP). Fourth ventricular lactate infusion exacerbates insulin-induced hypoglycemia (IIH) and IIH-associated patterns of DVC neuronal activation. We investigated the hypothesis that during glucoprivation, lactate regulates neuronal monocarboxylate and glucose transporter gene transcription in the DVC, and adjustments in these gene profiles are correlated with altered GK and SUR1 mRNA expression. We also examined whether caudal hindbrain lactate repletion alters the impact of hypoglycemia on substrate fuel uptake and metabolic sensing functions in other characterized metabolic monitoring sites, e.g., the ventromedial hypothalamic nucleus (VMH) and lateral hypothalamic area (LHA). qPCR was used to measure MCT2, GLUT3, GLUT4, GK, and SUR1 transcripts in the microdissected DVC, VMH, and LHA from groups of male rats treated by continuous infusion of aCSF or lactate into the caudal fourth ventricle (CV4), initiated prior to injection of Humulin R or saline. Blood glucose was decreased in response to insulin, a response that was significantly augmented by CV4 lactate infusion. IIH alone did not alter mean DVC MCT2, GLUT3, GLUT4, GK, or SUR1 mRNA levels, but these transcripts were increased in the lactate plus insulin group, relative to both euglycemic and aCSF-infused hypoglycemic rats. IIH decreased MCT2, GLUT3, and SUR1 gene profiles in the VMH; CV4 lactate infusion during IIH further diminished these transcripts, and suppressed GLUT4 and GK mRNA levels in this site. In LHA, IIH increased GLUT3 and SUR1 gene expression to an equal extent, with or without lactate, while GLUT4, MCT2, and GK mRNA levels were elevated only in response to lactate plus insulin. These studies show that caudal hindbrain-targeted delivery of exogenous lactate during IIH upregulates neuronal monocarboxylate and glucose transporter, GK, and SUR1 gene profiles in the DVC, and results in increased or decreased GLUT4 and GK mRNA in LHA and VMH, respectively. These data suggest that lactate and glucose utilization by DVC neurons may be enhanced in response to local lactate surfeit, alone or relative to glucose deficiency, and that increases in intracellular glucose and net energy yield may be correlated with elevated GK and SUR1 gene transcription, respectively, in local glucose sensing neurons. The results also imply that GLUT4- and GK-mediated glucose uptake and glucose sensing functions in the VMH and LHA may be reactive to DVC signaling of relative lactate abundance within the caudal hindbrain, and/or to physiological sequelae of this fuel augmentation, including amplified hypoglycemia.

Serotonin receptor involvement in effects of restraint on female rat lordosis behavior.

Ovariectomized Fischer (CDF-344) rats, with bilateral cannulae in the mediobasal hypothalamus (MBH) near the ventromedial nucleus of the hypothalamus (VMN), were used to test the hypothesis that serotonin receptors in the VMN contribute to the lordosis-inhibiting effects of mild restraint. Rats were hormonally primed with 10 microg estradiol benzoate (EB) followed 48 h later with sesame seed oil. Four to six hours later (during the dark portion of the light-dark cycle), rats were pretested for sexual behavior. Thereafter, they were infused with saline, 2 microg of the serotonin (5-HT) 2 receptor agonist, (+/-)-2,5-dimethoxy-4-iodophenyl-2-aminopropane HCl (DOI), or 1 microg of the 5-HT(1A) receptor antagonist, N-{2[4-(2-methoxyphenyl)-1-piperazinyl]ethyl}-N-(2-pyridinyl) cyclohexanecarboxamide trihydrochloride (WAY100635). After a 5 min restraint, rats were tested for sexual receptivity. Rats infused with saline showed a significant decline in lordosis behavior after restraint. Infusion with either DOI or WAY100635 attenuated these effects of restraint. These findings extend earlier observations that the lordosis-disruptive effects of mild restraint include activation of 5-HT(1A) receptors in the VMN and are the first to implicate VMN 5-HT(2) receptors in protection against mild restraint.

Androgen receptors are required for full masculinization of the ventromedial hypothalamus (VMH) in rats.

The ventromedial hypothalamus (VMH) is one of several sexually dimorphic nuclei that regulate mating behavior, and is rich in steroid hormone receptors and aromatase activity. We looked at the contribution of the androgen receptor (AR) to the volume of the VMH in rats by measuring each of the four subdivisions of the VMH in 90 day old male, female, and XY male rats carrying a mutant AR allele (tfm), which renders animals largely unresponsive to androgens. Confirming published reports, total VMH volume was greater in wild-type males than in females (P<0.01). The mean total volume of the VMH in TFM males was intermediate, but not significantly different from either females or males (Ps>0.10). The sex difference in VMH volume was primarily accounted for by the ventrolateral subdivision (VMHvl), which in both females and TFM males was significantly smaller than in wild-type males (Ps<0.005). There was no significant sex difference in the volume of the other three subdivisions of the VMH. Neuronal somata were larger in males than females in VMHvl, central VMH (VMHc) and the dorsomedial VMH (VMHdm), with TFM males having feminine neuronal somata in the VMHdm and VMHc. These data suggest that AR plays a role during sexual differentiation of the VMH, imparting its greatest effect in the VMHvl. ARs may regulate aromatase expression or activity to affect estrogen receptor activation, or may act independently of estrogen receptors to influence VMH morphology.

Anatomical distribution and biochemical characterization of the novel RFamide peptide 26RFa in the human hypothalamus and spinal cord.

26RFa is a novel RFamide peptide originally isolated in the amphibian brain. The 26RFa precursor has been subsequently characterized in various mammalian species but, until now, the anatomical distribution and the molecular forms of 26RFa produced in the CNS of mammals, in particular in human, are unknown. In the present study, we have investigated the localization and the biochemical characteristics of 26RFa-like immunoreactivity (LI) in two regions of the human CNS--the hypothalamus and the spinal cord. Immunohistochemical labeling using specific antibodies against human 26RFa and in situ hybridization histochemistry revealed that in the human hypothalamus 26RFa-expressing neurons are located in the paraventricular and ventromedial nuclei. In the spinal cord, 26RFa-expressing neurons were observed in the dorsal and lateral horns. Characterization of 26RFa-related peptides showed that two distinct molecular forms of 26RFa are present in the human hypothalamus and spinal cord, i.e. 26RFa and an N-terminally elongated form of 43 amino acids designated 43RFa. These data provide the first evidence that 26RFa and 43RFa are actually produced in the human CNS. The distribution of 26RF-LI suggests that 26RFa and/or 43RFa may modulate feeding, sexual behavior and transmission of nociceptive stimuli.

Steroid hormone mediation of limbic brain plasticity and aggression in free-living tree lizards, Urosaurus ornatus.

The neural mechanisms by which steroid hormones regulate aggression are unclear. Although testosterone and its metabolites are involved in both the regulation of aggression and the maintenance of neural morphology, it is unknown whether these changes are functionally related. We addressed the hypothesis that parallel changes in steroid levels and brain volumes are involved in the regulation of adult aggression. We examined the relationships between seasonal hormone changes, aggressive behavior, and the volumes of limbic brain regions in free-living male and female tree lizards (Urosaurus ornatus). The brain nuclei that we examined included the lateral septum (LS), preoptic area (POA), amygdala (AMY), and ventromedial hypothalamus (VMH). We showed that the volumes of the POA and AMY in males and the POA in females vary with season. However, reproductive state (and thus hormonal state) was incompletely predictive of these seasonal changes in males and completely unrelated to changes in females. We also detected male-biased dimorphisms in volume of the POA, AMY, and a dorsolateral subnucleus of the VMH but did not detect a dimorphism between alternate male morphological phenotypes. Finally, we showed that circulating testosterone levels were higher in males exhibiting higher frequency and intensity of aggressive display to a conspecific, though brain nucleus volumes were unrelated to behavior. Our findings fail to support our hypothesis and suggest instead that plasma testosterone level covaries with aggression level and in a limited capacity with brain nucleus volumes but that these are largely unrelated relationships.

Use of laser-capture microdissection for the identification of marker genes for the ventromedial hypothalamic nucleus.

The ventromedial hypothalamic nucleus (VMH) plays an important role in the control of feeding and energy homeostasis. In contrast to other hypothalamic nuclei that are also known to regulate energy balance, there is a paucity of nucleus-specific marker genes for the VMH, limiting the application of molecular approaches for analyzing VMH information processing, function, and circuitry. Here, we report the use of laser-capture microdissection to isolate a set of cDNAs that are enriched in the VMH relative to two adjacent hypothalamic nuclei, the arcuate and dorsomedial hypothalamus. The relative expression levels of nine of the 12 most robustly expressed VMH-enriched genes were confirmed by real-time PCR analysis using separate RNAs from these three nuclei. Three of these VMH-enriched genes were further characterized by in situ hybridization histochemistry, including pituitary adenylate cyclase activating polypeptide, cerebellin 1, and an expressed sequence tag named LBH2. Finally, to test whether some of these genes were coordinately regulated, we monitored their expression in steroidogenic factor 1 (SF-1) knock-out mice. SF-1 is a transcription factor that controls the development of the VMH. The RNA levels for four of these genes were reduced in these knock-out animals, further suggesting that they are direct or indirect targets of this orphan nuclear receptor. The VMH-enriched genes identified here provide a basis for a functional analysis of VMH neuronal subpopulations via the use of bacterial artificial chromosome transgenics and related technologies. These results also demonstrate the utility of laser-capture microdissection coupled with microarray technology to identify nucleus-specific transcriptional networks.

Estrogen modulates the sexually dimorphic synaptic connectivity of the ventromedial nucleus.

Neurons in the ventrolateral division of the hypothalamic ventromedial nucleus (VMNvl) display a remarkable estrogen-dependent functional and structural plasticity, which is likely to be mediated, in part at least, by neuronal afferents. The present study was designed to determine whether the number of synapses per neuron and the size of individual synapses in the VMNvl vary across the estrus cycle and, also, whether they differ between the sexes. To accomplish this, the VMNvl of adult female rats at proestrus or diestrus day 1 and of age-matched male rats was analyzed using electron microscopy. We found that a single VMNvl neuron receives around 7,000 synapses during diestrus and approximately 10,000 during proestrus. This estrus cycle-related variation is accounted for by increases in the number of all types of synapses. In males, the number of synapses received by each VMNvl neuron is similar to that of diestrus rats (approximately 7,500). However, in males the number of axodendritic and axospinous synapses is smaller than in proestrus rats, whereas the number of axosomatic synapses is higher than in diestrus rats. In addition, we found that the size of the postsynaptic densities of axospinous and axosomatic synapses is consistently larger in males than in females. Our results show that the synaptic organization of the VMNvl is sexually dimorphic, with females having more dendritic synapses and males more somatic synapses. They also show that the synaptic plasticity induced by estrogen in the VMNvl is characterized by changes in the number, but not the size, of the synapses.

Cyto- and chemoarchitecture of the hypothalamus of a wallaby ( Macropus eugenii) with special emphasis on oxytocin and vasopressinergic neurons.

We have studied the organization of the hypothalamus in an Australian diprotodontid metatherian mammal, the wallaby ( Macropus eugenii), using cytoarchitectural, histochemical and immunohistochemical techniques. Coronal sections of adult brains were processed for Nissl staining, histochemical reactivity (cytochrome oxidase, nicotinamide adenine dinucleotide phosphate diaphorase and acetylcholinesterase) and immunohistochemistry (antibodies to tyrosine hydroxylase, calbindin, calretinin, non-phosphorylated neurofilament protein, oxytocin and vasopressin). The distribution of immunoreactive neurons for these substances was mapped with the aid of a computer-linked microscope. In general, the wallaby hypothalamus showed a similar nuclear organization to that seen in rodents. The paraventricular nucleus could be divided into several subdivisions based on the different cellular parcellation, similar to that described in rodents. The ventromedial hypothalamic nucleus had cell-sparse dorsomedial and cell-dense ventrolateral subdivisions as seen in eutheria, suggesting a similar functional compartmentalization in all theria. The positions of tyrosine hydroxylase-positive neurons in the wallaby hypothalamus were also similar to those in eutheria. Oxytocin and vasopressinergic neurons were found in all the same major nuclear groups as seen in eutheria, although a nucleus circularis could not be identified. The general similarities between wallaby and eutherian hypothalamus indicate that the basic chemo- and cytoarchitectural features of the hypothalamus are common to eutheria and metatheria and validate the use of the wallaby as a mammalian model of wide applicability in investigations of hypothalamic functional development.

Fasting increases gene expressions of uncoupling proteins and peroxisome proliferator-activated receptor-gamma in brown adipose tissue of ventromedial hypothalamus-lesioned rats.

Uncoupling proteins (UCPs) are supposed to be involved in diet-induced thermogenesis. Their activities are usually elevated by feeding and reduced by fasting in normal animals. To investigate whether fasting affects the expression of UCPs mRNA in brown adipose tissue (BAT) of bilateral ventromedial hypothalamus (VMH)-lesioned rats, we determined the gene expression of UCP1, UCP2 or UCP3 in BAT of VMH-lesioned rats and examined oxygen consumption in these rats under fed or 48-h fasted conditions. Northern blotting revealed no difference in the expression of UCPs mRNA in BAT between VMH-lesioned and sham-operated rats under the fed condition, however, expressions were increased markedly in BAT of VMH-lesioned rats under the fasted condition. Under the fed condition, no difference in oxygen consumption was observed between VMH-lesioned and sham-operated rats. Under the fasted condition, oxygen consumption decreased in both rats, however, it decreased in VMH-lesioned less than in sham operated rats. To explore the mechanism that fasting elevated BAT UCPs mRNA in VMH-lesioned rats, we measured peroxisome proliferator-activated receptor (PPAR)-gamma mRNA and protein in BAT, because PPAR-gamma agonist can elevate UCPs mRNA levels in BAT. Under the fed condition, no differences in the expression of PPAR-gamma mRNA and protein content were observed between in BAT of VMH-lesioned and sham-operated rats. Under the fasted condition, however, both increased in BAT of VMH-lesioned rats. These results suggest that VMH-lesions enhance the gene expression of UCPs in BAT under long-term fasting as a defensive reaction to inhibit the reduction of body temperature through an increase in PPAR-gamma activity.

Fos-like immunoreactivity in brain regions of domestic rams following exposure to rams or ewes.

Limbic and basal forebrain-hypothalamic regions from male sheep differing in sexual performance were quantified for fos-like immunoreactivity. Rams classified as high-sexually performing (HP), low-sexually performing (LP), and male-oriented (MO) received noncontact sensory stimulation from either ewes in estrus (HP, n=5; LP, n=4; MO, n=4) or other males (HP, n=5; LP, n=4; MO, n=5) for a 4-h period on each of 3 consecutive days. Following exposure to stimulus animals on the third day, rams were euthanized and their brains were perfused with a 1% paraformaldehyde/1.5% glutaraldehyde solution and sections were analyzed for fos-like immunoreactivity. Brain regions analyzed were the medial amygdala (meAMY), medial preoptic area (mPOA), bed nucleus of the stria terminalis (BNST), and ventromedial hypothalamic nucleus (VMH). Fos-like immunoreactivity differed between groups in the mPOA and BNST but not in the meAMY or VMH. LP rams exposed to estrous ewes had more (P<.05) neurons staining positive for fos and fos-related antigens (FRA) in the mPOA and BNST than LP rams exposed to other rams or MO rams exposed to either estrous ewes or other rams. Numbers of neurons staining positive for FRA in the mPOA and BNST of LP rams exposed to estrous ewes, however, were not different (P>.05) from HP rams exposed to either estrous ewes or other rams. The similar fos-like immunoreactivity in areas important for the display of sexual behavior in HP and LP rams may reflect similar sensory input in these two groups of rams; however, LP rams, in contrast to HP rams, do not appear to respond similarly to the same sensory stimulus.