Anterior Hypothalamic Region Research Articles

VX-765 Alleviates Circadian Rhythm Disorder in a Rodent Model of Traumatic Brain Injury Plus Hemorrhagic Shock and Resuscitation.

Severe traumatic brain injury (TBI) can result in persistent complications, including circadian rhythm disorder, that substantially affect not only the injured people, but also the mood and social interactions with the family and the community. Pyroptosis in GFAP-positive astrocytes plays a vital role in inflammatory changes post-TBI. We determined whether VX-765, a low molecular weight caspase-1 inhibitor, has potential therapeutic value against astrocytic inflammation and pyroptosis in a rodent model of TBI plus hemorrhagic shock and resuscitation (HSR). A weight-drop plus bleeding and refusion model was used to establish traumatic exposure in rats. VX-765 (50mg/kg) was injected via the femoral vein after resuscitation. Wheel-running activity was assessed, brain magnetic resonance images were evaluated, the expression of pyroptosis-associated molecules including cleaved caspase-1, gasdermin D (GSDMD), and interleukin-18 (IL-18) in astrocytes in the region of anterior hypothalamus, were explored 30days post-trauma. VX-765-treated rats had significant improvement in circadian rhythm disorder, decreased mean diffusivity (MD) and mean kurtosis (MK), increased fractional anisotropy (FA), an elevated number and branches of astrocytes, and lower cleaved caspase-1, GSDMD, and IL-18 expression in astrocytes than TBI + HSR-treated rats. These results demonstrated that inhibition of pyroptosis-associated astrocytic activations in the anterior hypothalamus using VX-765 may ameliorate circadian rhythm disorder after trauma. In conclusion, we suggest that interventions targeting caspase-1-induced astrocytic pyroptosis by VX-765 are promising strategies to alleviate circadian rhythm disorder post-TBI.

Impact of the circadian clock on cardiovascular physiology and pathophysiology

Circadian rhythms are the innate approximately 24-hour oscillations that orchestrate a multitude of physiological processes encompassing sleep-wake cycles, hormone secretion, thermoregulation, metabolism, and behavioral patterns. Also, those rhythms are intrinsically governed by the suprachiasmatic nucleus (SCN) ensconced in the anterior hypothalamic region. The SCN, synchronized during daylight hours receives and transmits diverse bodily signals to calibrate physiological dynamics, and imbue disturbances with potential repercussions reverberating across a plethora of physiological and behavioral domains. Previous studies have proven that the diurnal changes in the circadian rhythm multiple myocardial processes and have a critical impact on the physiology and pathophysiology of the cardiovascular system. This comprehensive review emphasizes the profound influence of circadian rhythms, orchestrated by the suprachiasmatic nucleus, on cardiovascular health. It reveals that disruptions in circadian rhythms impact blood pressure, myocardial function, platelet responsiveness, and the autonomic nervous system, all of which are critical for cardiovascular well-being. The significance of this research lies in its potential to guide interventions for cardiovascular disease prevention and management, particularly in the context of modern challenges like shift work. Understanding this intricate relationship is crucial for safeguarding cardiovascular health in our fast-paced world.

Changes in pituitary gonadotropin subunits and hypothalamic Kiss-1 gene expression by administration of sex steroids in ovary-intact female rats.

We examined how the sex steroids influence the synthesis of gonadotropins. The effects of sex steroids estradiol (E2), progesterone (P4), and dihydrotestosterone (DHT) in pituitary gonadotroph cell model (LβT2 cells) in vitro and ovary-intact rats in vivo were examined. The effects of sex steroids on Kiss1 gene expression in the hypothalamus were also examined in ovary-intact rats. In LβT2 cells, E2 increased common glycoprotein alpha (Cga) and luteinizing hormone beta (Lhb) subunit promoter activity as well as their mRNA expression. Although gonadotropin subunit promoter activity was not modulated by P4, Cga and Lhb mRNA expression was increased by P4. DHT inhibited Cga and Lhb mRNA expression with a concomitant decrease in their promoter activity. During the 2-week administration of exogenous E2 to ovary-intact rats, the estrous cycle determined by vaginal smears was disrupted. P4 or DHT administration completely eliminated the estrous cycle. Protein expression of all three gonadotropin subunits within the pituitary gland was inhibited by E2 or P4 treatment in vivo; however, DHT reduced Cga expression but did not modulate Lhb or follicle-stimulating hormone beta subunit expression. E2 administration significantly repressed Kiss1 mRNA expression in a posterior hypothalamic region that included the arcuate nucleus. P4 and DHT did not modulate Kiss1 mRNA expression in this region. In contrast, P4 administration significantly inhibited Kiss1 mRNA expression in the anterior region of the hypothalamus that included the anteroventral periventricular nucleus. The expression of gonadotropin-releasing hormone (Gnrh) mRNA in the anterior hypothalamic region, where the preoptic area is located, appeared to be decreased by treatment with E2 and P4. Our findings suggest that sex steroids have different effects in the hypothalamus and pituitary gland.

Fifteen-minute consultation: Paediatric night sweats—when to reassure and when to investigate

3-year-old Susie has been referred to the medical paediatric clinic by her general practitioner with sweating at night; her parents change Susie’s pillowcase whilst she is asleep at around 11pm...

Predicting the location of the preoptic and anterior hypothalamic region by visualizing the thermoregulatory center on fMRI in craniopharyngioma using cold and warm stimuli.

Hypothalamic nuclei in the preoptic and anterior hypothalamic region (POAH) are critically involved in thermoregulation and neuroendocrine regulation and can be displaced by craniopharyngiomas (CPs). We aimed to locate the POAH by visualizing hypothalamic thermoregulation through task-related functional magnetic resonance imaging (fMRI) to guide hypothalamus protection intraoperatively. Nine adult healthy volunteers (HVs) and thirty-two adult primary CP patients underwent task-related fMRI for POAH localization by warm (60° C) and cold (0° C) cutaneous thermoreceptor stimulation. Approach selection and intraoperative POAH protection were performed based on preoperative POAH localization. In all HVs and patients, significant single positive blood oxygen level-dependent (BOLD) signal changes were located in the POAH. The BOLD activity was significantly greater for cold (P=0.03) and warm (P=0.03) stimuli in patients than in HVs. Gross total resection and near-total resection were achieved in 28 (87.5%) and 4 (12.5%) patients, respectively. New-onset diabetes insipidus and new-onset hypopituitarism occurred in 6 patients (18.8%) and 10 patients (31.3%), respectively. Our findings suggest that cutaneous thermoreceptor stimulation could accurately activate the hypothalamic thermoregulatory center and allow POAH localization through task-related fMRI. Preoperative POAH localization could help neurosurgeons protect hypothalamic function intraoperatively. The CP patients were more sensitive to thermal stimulation.

The Lateral Preoptic Area and Its Projection to the VTA Regulate VTA Activity and Drive Complex Reward Behaviors

The ventral tegmental area (VTA) underlies motivation and reinforcement of natural rewards. The lateral preoptic area (LPO) is an anterior hypothalamic brain region that sends direct projections to the VTA and to other brain structures known to regulate VTA activity. Here, we investigated the functional connection between the LPO and subpopulations of VTA neurons and explored the reinforcing and valence qualities of the LPO in rats. We found that the LPO and the LPO→VTA pathway inhibit the activity of VTA GABA neurons and have mixed effects on VTA dopamine neurons. Furthermore, we found that the LPO supports operant responding but drives avoidance, and we explored the apparent discrepancy between these two results. Finally, using fiber photometry, we show that the LPO signals aversive events but not rewarding events. Together, our findings demonstrate that the LPO modulates the activity of the VTA and drives motivated behavior and represents an overlooked modulator of reinforcement.

Does the Apparent Diffusion Coefficient Value Predict Permanent Cerebral Ischemia/Reperfusion Injury in Rats?

Variation in tissue damage after cerebral ischemia/reperfusion (I/R) can cause uncertainty in stroke-related studies, which can be reduced if the damage can be predicted early after ischemia by measuring the apparent diffusion coefficient (ADC). We investigated whether ADC measurement in the acute phase can predict permanent cerebral I/R injury. The middle cerebral artery occlusion model was established using the intraluminal suture method to induce 60 minutes of ischemia followed by reperfusion in rats. T2-weighted images and diffusion-weighted images were obtained at 30 minutes and 24 hours after ischemia. Neuronal cell survival was assessed by neuronal nuclei (NeuN) immunofluorescence staining. The correlation between relative ADC (rADC) values at 30 minutes and I/R injury at 24 hours after ischemia was analyzed. Magnetic resonance imaging results were confirmed by histologic analysis. The correlation between rADC values at 30 minutes and 24 hours was strong in the ischemic core and peri-infarct region but moderate in the anterior choroidal and hypothalamic region. Histologic analysis revealed that the correlation between rADC values at 30 minutes and the number of NeuN-positive cells at 24 hours was strong in the ischemic core and peri-infarct region but moderate in the anterior choroidal and hypothalamic region. Furthermore, there was a strong positive correlation between the sum of rADC values of three regions at 30 minutes and the infarct volume at 24 hours. ADC measurement in the acute phase can predict permanent cerebral I/R injury and provide important information for the evaluation of ischemic stroke.

Area-specific analysis of the distribution of hypothalamic neurons projecting to the rat ventral tegmental area, with special reference to the GABAergic and glutamatergic efferents

The ventral tegmental area (VTA) is a main regulator of reward and integrates a wide scale of hormonal and neuronal information. Feeding-, energy expenditure-, stress, adaptation- and reproduction-related hypothalamic signals are processed in the VTA and influence the reward processes. However, the neuroanatomical origin and chemical phenotype of neurons mediating these signals to the VTA have not been fully characterized. In this study we have systematically mapped hypothalamic neurons that project to the VTA using the retrograde tracer Choleratoxin B subunit (CTB) and analyzed their putative gamma-aminobutyric acid (GABA) and/or glutamate character with in situ hybridization in male rats. 23.93 ± 3.91% of hypothalamic neurons projecting to the VTA was found in preoptic and 76.27 ± 4.88% in anterior, tuberal and mammillary hypothalamic regions. Nearly half of the retrogradely-labeled neurons in the preoptic, and more than one third in the anterior, tuberal and mammillary hypothalamus appeared in medially located regions. The analyses of vesicular glutamate transporter 2 (VGLUT2) and glutamate decarboxylase 65 (GAD65) mRNA expression revealed both amino acid markers in different subsets of retrogradely-labeled hypothalamic neurons, typically with the predominance of the glutamatergic marker VGLUT2. About one tenth of CTB-IR neurons were GAD65-positive even in hypothalamic nuclei expressing primarily VGLUT2. Some regions were populated mostly by GAD65 mRNA-containing retrogradely-labeled neurons. These included the perifornical part of the lateral hypothalamus where 58.63 ± 19.04% of CTB-IR neurons were GABAergic. These results indicate that both the medial and lateral nuclear compartments of the hypothalamus provide substantial input to the VTA. Furthermore, colocalization studies revealed that these projections not only use glutamate but also GABA for neurotransmission. These GABAergic afferents may underlie important inhibitory mechanism to fine-tune the reward value of specific signals in the VTA.

Deep-brain photoreceptors (DBPs) involved in the photoperiodic gonadal response in an avian species, Gallus gallus

Three primitive photoreceptors [melanopsin (Opn4), neuropsin/opsin5 (Opn5) and vertebrate ancient opsin (VAOpn)] were reported as possible avian deep-brain photoreceptors (DBPs) involved in the perception of photoperiodic information affecting the onset and development of reproduction. The objective of this study was to determine the effect of long-day photostimulation and/or sulfamethazine treatment (SMZ, a compound known to advance light-induced testes development) on gene expression of DBPs and key hypothalamic and pituitary genes involved in avian reproductive function. Two-week old chicks were randomly selected into four experimental groups: short-day control (SC, LD8:16), short-day+SMZ (SS, LD8:16, 0.2% diet SMZ), long-day control (LC, LD16:8), and long-day+SMZ (LS, LD16:8, 0.2% diet SMZ). Birds were sampled on days 3, 7, and 28 after initiation of a long-day photoperiod and/or SMZ dietary treatments. Three brain regions [septal–preoptic, anterior hypothalamic (SepPre/Ant-Hypo) region, mid-hypothalamic (Mid-Hypo) region, posterior-hypothalamic (Post-Hypo) region], and anterior pituitary gland were dissected. Using quantitative real-time RT-PCR, we determined changes of expression levels of genes in distinct brain regions; Opn4 and Opn5 in SepPre/Ant-Hypo and Post-Hypo regions and, VAOpn in the Mid-Hypo region. Long-day treatment resulted in a significantly elevated testes weight on days 7 and 28 compared to controls, and SMZ augmented testes weight in both short- and long-day treatment after day 7 (P<0.05). Long-day photoperiodic treatment on the third day unexpectedly induced a large 8.4-fold increase of VAOpn expression in the Mid-Hypo region, a 15.4-fold increase of Opn4 and a 97.8-fold increase of Opn5 gene expression in the Post-Hypo region compared to SC birds (P<0.01). In contrast, on days 7 and 28, gene expression of the three DBPs was barely detectable. LC group showed a significant increase in GnRH-1 and TRH mRNA in the Mid-Hypo compared to SC on day 3. Pituitary LHβ and FSHβ mRNA were significantly elevated in LC and LS groups compared to SC on days 3 and 7 (P<0.05). On days 3 and 7, TSHβ mRNA level was significantly elevated by long-day treatment compared to the SC groups (P<0.05). Results suggest that long-day photoperiodic activation of DBPs is robust, transient, and temporally related with neuroendocrine genes involved in reproductive function. Additionally, results indicate that two subsets of GnRH-1 neurons exist based upon significantly different gene expression from long-day photostimulation and long-day plus SMZ administration. Taken together, the data indicate that within 3days of a long-day photoperiod, an eminent activation of all three types of DBPs might be involved in priming the neuroendocrine system to activate reproductive function in birds.

Different Critical Perinatal Periods and Hypothalamic Sites of Oestradiol Action in the Defeminisation of Luteinising Hormone Surge and Lordosis Capacity in the Rat

Female rats show a gonadotrophin-releasing hormone (GnRH)/luteinising hormone (LH) surge in the presence of a preovulatory level of oestrogen, whereas males do not because of brain defeminisation during the developmental period by perinatal oestrogen converted from androgen. The present study aimed to identify the site(s) of oestrogen action and the critical period for defeminising the mechanism regulating the GnRH/LH surge. Animals given perinatal treatments, such as steroidal manipulations, brain local implantation of oestradiol (E(2) ) or administration of an NMDA antagonist, were examined for their ability to show an E(2) -induced LH surge at adulthood. Lordosis behaviour was examined to compare the mechanisms defeminising the GnRH/LH surge and sexual behaviour. A single s.c. oestradiol-benzoate administration on either the day before birth (E21), the day of birth (D0) or day 5 (D5) postpartum completely abolished the E(2) -induced LH surge at adulthood in female rats, although the same treatment did not inhibit lordosis. Perinatal castration on E21 or D0 partially rescued the E2-induced LH surge in genetically male rats, whereas castration from E21 to D5 totally rescued lordosis. Neonatal E(2) implantation in the anterior hypothalamus including the anteroventral periventricular nucleus (AVPV)/preoptic area (POA) abolished the E(2) -induced LH surge in female rats, whereas E(2) implantation in the mid and posterior hypothalamic regions had no inhibitory effect on the LH surge. Lordosis was not affected by neonatal E(2) implantation in any hypothalamic regions. In male rats, neonatal NMDA antagonist treatment rescued lordosis but not the LH surge. Taken together, these results suggest that an anterior hypothalamic region such as the AVPV/POA region is a perinatal site of oestrogen action where the GnRH/LH regulating system is defeminised to abolish the oestrogen-induced surge. The mechanism for defeminisation of the GnRH/LH surge system might be different from that of sexual behaviour, in terms of the site(s) of oestrogen action and critical period, as well as the neurotransmitter system involved.

Cellular location and major terminal networks of the orexinergic system in the brains of five microchiropteran species

The present study describes the distribution of Orexin-A immunoreactive cell bodies and terminal networks in the brains of five microchiropteran species. Given the specialized flight and echolocation abilities of the microchiropterans it was of interest to examine if any specific differences in a generally phylogenetically homogenous neural system could be found. The orexinergic neurons have been found within the hypothalamus of all species studied, and were represented by a large cluster that spanned the anterior, dorsomedial, perifornical and lateral hypothalamic regions, with a smaller cluster extending into the region of the medial zona incerta. Evidence for orexinergic neurons in the ventrolateral hypothalamus adjacent to the optic tract was not observed in any microchiropteran species. The terminal networks of the orexinergic neurons conformed to that previously reported in a range of mammalian species, with dense terminal networks being found in the hypothalamus, cholinergic pedunculopontine and laterodorsal tegemental nuclei, the noradrenergic locus coeruleus complex, all serotonergic nuclei, the paraventricular nuclei of the epithalamus and adjacent to the habenular nuclei. Thus, apart from the lack of neurons in the ventrolateral hypothalamus, the orexinergic system of the microchiropterans appears typically mammalian.

Adolescent exposure to anabolic/androgenic steroids and the neurobiology of offensive aggression: A hypothalamic neural model based on findings in pubertal Syrian hamsters

Considerable public attention has been focused on the issue of youth violence, particularly that associated with drug use. It is documented that anabolic steroid use by teenagers is associated with a higher incidence of aggressive behavior and serious violence, yet little is known about how these drugs produce the aggressive phenotype. Here we discuss work from our laboratory on the relationship between the development and activity of select neurotransmitter systems in the anterior hypothalamus and anabolic steroid-induced offensive aggression using pubertal male Syrian hamsters ( Mesocricetus auratus) as an adolescent animal model, with the express goal of synthesizing these data into an cogent neural model of the developmental adaptations that may underlie anabolic steroid-induced aggressive behavior. Notably, alterations in each of the neural systems identified as important components of the anabolic steroid-induced aggressive response occurred in a sub-division of the anterior hypothalamic brain region we identified as the hamster equivalent of the latero-anterior hypothalamus, indicating that this sub-region of the hypothalamus is an important site of convergence for anabolic steroid-induced neural adaptations that precipitate offensive aggression. Based on these findings we present in this review a neural model to explain the neurochemical regulation of anabolic steroid-induced offensive aggression showing the hypothetical interaction between the arginine vasopressin, serotonin, dopamine, γ-aminobutyric acid, and glutamate neural systems in the anterior hypothalamic brain region.

Changes in the Expression of с-fos and NADPH-Diaphorase Activity in Rat Hippocampal Structures Related to Food Deprivation and Realization of Operant Food-Procuring Movements

The expression of early c-fos gene (marker of neuronal activation) and NADPH-diaphorase reactivity (NADPH-dr) was studied in various hypothalamic structures of rats in the norm, in the state of starvation, and after realization of long-lasting (repeated 4 to 12 times per minute for 30 min) motivated stereotyped food-procuring forelimb movements. In rats in the starving state, as compared with the control, the densities (number of units within a 200 × 200 μm 2 test area of a 40-μm-thick slice) of Fos-immunoreactive (Fos-ir) neurons in the parvicellular part of the paraventricular nucleus (Ра), supraoptic (SO), and medial preoptic (МРО) nuclei, anterior hypothalamic region (АН), and lateral hypothalamic nucleus (LH) were significantly greater (Р < 0.05) than in the control. In the dorsomedial (DMD) and ventromedial (VMHD) hypothalamic nuclei, this index did not differ from control values. After the performance of intense unilateral operant movements, higher densities of labeled neurons (as compared with that in control and starving animals) were observed in the PаAP, SO, МРО, and DMD, while smaller densities were observed in the LH and VMH. NADPH-dr neurons (i.e., NO synthase-containing cells) were observed in many hypothalamic nuclei; the maximum density of such NO-generating neurons was found in the Pa, SO, MPO, and DMD. The overwhelming majority of Fos-ir and NADPH-dr neurons in neurons was observed after realization of stereotyped food-procuring movements in the Ра and SO. This specificity of changes in the number of Fos-irand NADPH-dr neurons in the hypothalamic nuclei reflects, perhaps, the involvement of these structures in the control of autonomic functions in the course of realization of operant reflexes and adaptation of the function of the cardiovascular system to the corresponding intense physical and emotional loading.

Regulation of the neuronal norepinephrine transporter by endothelin-1 and -3 in the rat anterior and posterior hypothalamus

We previously reported that endothelin-1 and endothelin-3 modulate norepinephrine neuronal release and tyrosine hydroxylase activity and expression in the hypothalamus. In the present study we sought to establish the role of endothelin-1 and -3 in the regulation of norepinephrine uptake in the anterior and posterior hypothalamus. Results showed that in the anterior hypothalamus endothelin-3 increased neuronal norepinephrine uptake whereas endothelin-1 decreased it. Conversely, in the posterior hypothalamic region both endothelins diminished the neuronal uptake of the amine. Endothelins response was concentration dependent and maintained at all studied times. Endothelins also modified the kinetic and internalization of the NE neuronal transporter. In the anterior hypothalamic region endothelin-3 increased the V max and the B max whereas endothelin-1 decreased them. However, in the posterior hypothalamic region both endothelins diminished the V max as well as B max. Neither endothelin-1 nor endothelin-3 modified neuronal norepinephrine transporter K d in the studied hypothalamic regions. These findings support that in the posterior hypothalamic region both endothelins diminished neuronal norepinephrine transporter activity by reducing the amine transporter expression on the plasmatic membrane. Conversely, in the anterior hypothalamic region endothelin-3 enhanced neuronal norepinephrine transporter activity by increasing the expression of the transporter on the presynaptic membrane, whereas endothelin-1 induced the opposite effect. Present results permit us to conclude that both endothelins play an important role in the regulation of norepinephrine neurotransmission at the presynaptic nerve endings in the hypothalamus.

Reproductive functions and hypothalamic catecholamines in response to the soil fumigant metam sodium: Adaptations to extended exposures

Metam sodium (MS) is a soil fumigant and Category II pesticide with a relatively low toxicity in mammals. Previous data have shown an ability to impair reproductive mechanisms in ovariectomized, estradiol-primed rats. A single i.p. injection blocked the luteinizing hormone (LH) surge that in gonadal-intact females initiates the final stages of follicular and oocytic maturation and serves as the trigger for ovulation. The effect paralleled a fall in hypothalamic norepinephrine (NE) and rise in hypothalamic dopamine (DA) that was likely due to a suppression in dopamine- β-hydroxylase activity. In addition to determining the influence on catecholamine (CA) concentrations from a single oral exposure to MS, the present study explored effects of longer, 3-week treatments on estrous cyclicity, the LH surge, ovulation and hypothalamic CAs. Normally cycling 90 d S–D rats were administered MS (0–200 mg/kg/d, oral) and cyclicity was monitored daily. At the end of the 3rd week, proestrous blood was sampled over the afternoon from regular 4-day cyclers for a determination of LH. These animals were then killed on the following day of estrus (treatment days 21–26) for oocyte retrieval and assessment of hypothalamic CAs. Results showed that shortly after treatment began there occurred a dose-related period of persistent diestrus that typically lasted 8–16 d before regular cycles were reinstated. After 3 weeks, no effects were seen on the magnitude/timing of the LH surge or ovulated oocyte numbers. Anterior and posterior hypothalamic NE and DA were not significantly different from controls, although DA turnover (reflected by the ratio of DOPAC {3,4-dihydroxy-phenylacetic acid} to DA) in both anterior hypothalamic and caudate regions was decreased at all dosages. The data indicate that a 3 week oral exposure to MS induced an initial period of extended diestrus before the resumption of apparently normal reproductive activity, with previously reported CA alterations (apart from a persistent alteration in the DOPAC/DA ratio) being normalized by the end of dosing.

Immunohistochemical evidence for an endocrine/paracrine role for ghrelin in the reproductive tissues of sheep

BackgroundThe gut hormone, ghrelin, is involved in the neuroendocrine and metabolic responses to hunger. In monogastric species, circulating ghrelin levels show clear meal-related and body weight-related changes. The pattern of secretion and its role in ruminant species is less clear. Ghrelin acts via growth hormone secretagogue receptors (GHSR-1a) to alter food intake, fat utilization, and cellular proliferation. There is also evidence that ghrelin is involved in reproductive function. In the present study we used immunohistochemistry to investigate the presence of ghrelin and GHSR-1a in sheep reproductive tissues. In addition, we examined whether ghrelin and GHSR-1a protein expression is developmentally regulated in the adult and fetal ovine testis, and whether there is an association with markers of cellular proliferation, i.e. stem cell factor (SCF) and proliferating cell nuclear antigen (PCNA).MethodsAntibodies raised against ghrelin and its functional receptor, GHSR-type 1a, were used in standard immunohistochemical protocols on various reproductive tissues collected from adult and fetal sheep. GHSR-1a mRNA presence was also confirmed by in situ hybridisation. SCF and PCNA immunoexpression was investigated in fetal testicular samples. Adult and fetal testicular immunostaining for ghrelin, GHSR-1a, SCF and PCNA was analysed using computer-aided image analysis. Image analysis data were subjected to one-way ANOVA, with differences in immunostaining between time-points determined by Fisher's least significant difference.ResultsIn adult sheep tissue, ghrelin and GHSR-1a immunostaining was detected in the stomach (abomasum), anterior pituitary gland, testis, ovary, and hypothalamic and hindbrain regions of the brain. In the adult testis, there was a significant effect of season (photoperiod) on the level of immunostaining for ghrelin (p < 0.01) and GHSR-1a (p < 0.05). In the fetal sheep testis, there was a significant effect of gestational age on the level of immunostaining for ghrelin (p < 0.001), GHSR-1a (p < 0.05), SCF (p < 0.05) and PCNA (p < 0.01).ConclusionEvidence is presented for the presence of ghrelin and its receptor in various reproductive tissues of the adult and fetal sheep. In addition, the data indicate that testicular expression of ghrelin and its receptor is physiologically regulated in the adult and developmentally regulated in the fetus. Therefore, the ghrelin ligand/receptor system may have a role (endocrine and/or paracrine) in the development (cellular proliferation) and function of the reproductive axis of the sheep.

5-hydroxytryptamine release in the anterior hypothalamic and the hippocampal areas of cholestatic rats

Cholestasis contributes to the genesis of fatigue through several mechanisms. Among these mechanisms, affected serotonergic neurotransmission is important in the pathogenesis of central fatigue. Previously, elevated levels of 5-hydroxyindole acetic acid (5-HIAA), the metabolite of 5-hydroxytryptamine (5-HT) and increased 5-HT 2 receptor density were demonstrated in the anterior hypothalamus and in the hippocampus of bile duct resected rats (BDR), respectively. The aim of this paper is to demonstrate evoked 5-HT release in selected brain regions like anterior hypothalamus and hippocampal CA1 regions of cholestatic rats using BDR rats as an experimental model for cholestasis. In this study, we analyzed the K + evoked 5-HT and its metabolite 5-HIAA levels by using HPLC with electrochemical detection in the microdialysis samples collected from anterior hypothalamic and hippocampal CA1 regions of sham-operated and BDR rats ( n = 6). The ratios of [5-HIAA] to [5-HT] following perfusion with 100 mM K + artificial cerebrospinal fluid was used for the comparison of the evoked release of 5-HT. Locomotor activity was used to assess the signs of cholestasis associated fatigue in the BDR rats. The vertical and horizontal activity counts within 15 min were found to be decreased in the BDR rats compared to sham-operated rats ( p < 0.05). Besides, the number of fecal boli (an index of emotionality) was also significantly fewer in the cholestatic rats ( p < 0.05). No significant difference between the sham-operated and the BDR rats was detected in the basal 5-HT and 5-HIAA levels of anterior hypothalamus. K + stimulation yielded a more profound increase in the [5-HIAA] / [5-HT] in the BDR rats ( p < 0.05). The basal levels of 5-HT in CA1 region of the BDR rats was found to be lower than that of sham-operated group ( p < 0.05), but no significant difference was observed in terms of evoked 5-HT release in both sham-operated and BDR rats. These findings imply the presence of affected serotonergic system in cholestasis.

Involvement of nitric oxide pathways in short term modulation of tyrosine hydroxylase activity by endothelins 1 and 3 in the rat anterior hypothalamus

The ability of endothelins 1 and 3 (ET-1 and ET-3) to reduce neuronal norepinephrine release through ETB receptor activation involving nitric oxide (NO) pathways in the rat anterior hypothalamus region (AHR) was previously reported. In the present work, we studied the effects of ET-1 and -3 on tyrosine hydroxylase (TH) activity and the possible involvement of NO pathways. Results showed that ET-1 and -3 (10 nM) diminished TH activity in AHR and this effect was blocked by a selective ETB receptor antagonist (100 nM BQ-788), but not by a ET A receptor antagonist (BQ-610). To confirm these results, 1 μM IRL-1620 (ET B agonist) reduced TH activity whereas 300 nM sarafotoxin S6b falled to modify it. N ω-Nitro- l-arginine methyl ester (10 μM), 7-nitroindazole (10 μM), 1 H-[1,2,4]oxadiazolo[4,3- a]quinoxalin-1-ona (10 μM), KT5823 (2 μM), inhibitors of nitric oxide synthase, neuronal nitric oxide synthase, NO-sensitive-guanylyl cyclase, and protein kinase G, respectively, did not modify the reduction of TH activity produced by ETs. In addition, both 100 μM sodium nitroprusside and 50 μM 8-bromoguanosine-3′,5′-cyclic monophosphate (NO donor and guanosine-3′,5′-cyclic monophosphate analog, respectively) diminished TH activity. Present results showed that ET-1 and ET-3 diminished TH activity through the activation of ET B receptors involving the NO/guanosine-3′,5′-cyclic monophosphate/protein kinase G pathway. Taken jointly present and previous results it can be concluded that both ETs play an important role as modulators of norepinephrine neurotransmission in the rat AHR.

Absence of the cholecystokinin-A receptor deteriorates homeostasis of body temperature in response to changes in ambient temperature.

The circadian rhythm of the body core temperature (T(c)) and the effects of changes in ambient temperatures on the homeostasis of T(c) in Otsuka Long Evans Tokushima Fatty (OLETF) rats, which are naturally occurring cholecystokinin (CCK)-A receptor (CCK-AR) gene knockout (-/-) rats, were examined. In addition, the peripheral responses to warming or cooling of the preoptic and anterior hypothalamic region (PO/AH) were determined. The circadian rhythm of T(c) in OLETF rats was similar to that in Long-Evans Tokushima (LETO) rats; this rhythm was characterized by a higher T(c) during the dark period and a lower T(c) during the light period. When the ambient temperature was changed within the limits of 0 degrees C to 30 degrees C, the changes in T(c) of LETO rats were associated with the changes in ambient temperature, whereas those in OLETF rats were dissociated from the temperature changes. The OLETF rats showed a large hysteresis. The peripheral responses to warming or cooling of PO/AH, including shivering of the neck muscle and changes in skin temperature of the tail and footpad, were similar in OLETF and LETO rats. To confirm the role of CCK-AR in the regulation of body temperature, the values of T(c) in the CCK-AR(-/-) mice were compared with those in CCK-B receptor (CCK-BR) (-/-), CCK-AR(-/-)BR(-/-), and wild-type mice. In the mice, the circadian rhythms of T(c) were the same, regardless of the genotype. Mice without CCK-AR showed larger hysteresis than mice with CCK-AR. From these results, we conclude that the lack of CCK-AR causes homeostasis of T(c) in rats and mice to deteriorate.

Biotinylated Dextran Amine as a Marker for Fetal Hypothalamic Homografts and Their Efferents

We have explored the use of biotinylated dextran amine (BDA) as a marker for labeling fetal brain grafts and their connections with the host. As a model system we used transplantation of the hamster suprachiasmatic nucleus, the site of an endogenous biological clock governing circadian rhythms. Similar transplants into arrhythmic hosts have been shown to restore behavioral function with a period specific to the donor. For locomotor rhythms, efferent connections are not necessary. For other responses, including endocrine rhythms, efferent connections may be necessary. In order to visualize homografts and their efferents, injections of BDA, an anterograde tracer, were made into the anterior hypothalamic (AH) region containing the SCN or into the dorsal cortex (CTX) of fetal hamster brains. The fetal AH or CTX was microdissected out and stereotaxically implanted into the third ventricle of intact, adult hamsters. After 2, 4, 8, or 12 weeks, hosts were sacrificed and their brains were processed for detection of BDA by either histochemistry or immunofluorescence. BDA intensely labeled graft neurons, their dendrites, and axons with minimal or no spread to the adjacent host brain. Labeled graft axons could be followed for long distances (>1 mm) into the host brain and graft-derived varicosities formed close contacts with host neurons. BDA-labeled graft neurons, located at the perimeter of the graft, also extended dendrite-like processes into the host parenchyma. We conclude that BDA is a useful marker for fetal homografts and their efferents for survival times of less than 2 months.