Ablation of Growth Hormone Receptor in GABAergic Neurons Leads to Increased Pulsatile Growth Hormone Secretion.

Growth hormone (GH) acts in several hypothalamic neuronal populations to modulate metabolism and the autoregulation of GH secretion via negative-feedback loops. However, few studies have investigated whether GH receptor (GHR) expression in specific neuronal populations is required for the homeostatic control of GH secretion and energy homeostasis. In the present study, we investigated the consequences of the specific GHR ablation in GABAergic (VGAT-expressing) or glutamatergic (VGLUT2-expressing) cells. GHR ablation in GABAergic neurons led to increased GH secretion, lean mass, and body growth in male and female mice. VGAT-specific GHR knockout (KO) male mice also showed increased serum insulin-like growth factor-1, hypothalamic Ghrh, and hepatic Igf1 messenger RNA levels. In contrast, normal GH secretion, but reduced lean body mass, was observed in mice carrying GHR ablation in glutamatergic neurons. GHR ablation in GABAergic cells increased weight loss and led to decreased blood glucose levels during food restriction, whereas VGLUT2-specific GHR KO mice showed blunted feeding response to 2-deoxy-D-glucose both in males and females, and increased relative food intake, oxygen consumption, and serum leptin levels in male mice. Of note, VGLUT2-cre female mice, independently of GHR ablation, exhibited a previously unreported phenotype of mild reduction in body weight without further metabolic alterations. The autoregulation of GH secretion via negative-feedback loops requires GHR expression in GABAergic cells. Furthermore, GHR ablation in GABAergic and glutamatergic neuronal populations leads to distinct metabolic alterations. These findings contribute to the understanding of the neuronal populations responsible for mediating the neuroendocrine and metabolic effects of GH.

Leptin Receptor Expression in GABAergic Cells is Not Sufficient to Normalize Metabolism and Reproduction in Mice.

Previous studies indicate that leptin receptor (LepR) expression in GABAergic neurons is necessary for the biological effects of leptin. However, it is not clear whether LepR expression only in GABAergic neurons is sufficient to prevent the metabolic and neuroendocrine imbalances caused by LepR deficiency. In the present study, we produced mice that express the LepR exclusively in GABAergic cells (LepRVGAT mice) and compared them with wild-type (LepR+/+) and LepR-deficient (LepRNull/Null) mice. Although LepRVGAT mice showed a pronounced reduction in body weight and fat mass, as compared with LepRNull/Null mice, male and female LepRVGAT mice exhibited an obese phenotype relative to LepR+/+ mice. Food intake was normalized in LepRVGAT mice; however, LepRVGAT mice still exhibited lower energy expenditure in both sexes and reduced ambulatory activity in the females, compared with LepR+/+ mice. The acute anorexigenic effect of leptin and hedonic feeding were normalized in LepRVGAT mice despite the hyperleptinemia they present. Although LepRVGAT mice showed improved glucose homeostasis compared with LepRNull/Null mice, both male and female LepRVGAT mice exhibited insulin resistance. In contrast, LepR expression only in GABAergic cells was sufficient to normalize the density of agouti-related peptide (AgRP) and α-MSH immunoreactive fibers in the paraventricular nucleus of the hypothalamus. However, LepRVGAT mice exhibited reproductive dysfunctions, including subfertility in males and alterations in the estrous cycle of females. Taken together, our findings indicate that LepR expression in GABAergic cells, although critical to the physiology of leptin, is insufficient to normalize several metabolic aspects and the reproductive function in mice.

Synaptic Vesicle Protein 2A Expression in Glutamatergic Terminals Is Associated with the Response to Levetiracetam Treatment.

Synaptic vesicle protein 2A (SV2A), the target of the antiepileptic drug levetiracetam (LEV), is expressed ubiquitously in all synaptic terminals. Its levels decrease in patients and animal models of epilepsy. Thus, changes in SV2A expression could be a critical factor in the response to LEV. Epilepsy is characterized by an imbalance between excitation and inhibition, hence SV2A levels in particular terminals could also influence the LEV response. SV2A expression was analyzed in the epileptic hippocampus of rats which responded or not to LEV, to clarify if changes in SV2A alone or together with glutamatergic or GABAergic markers may predict LEV resistance. Wistar rats were administered saline (control) or pilocarpine to induce epilepsy. These groups were subdivided into untreated or LEV-treated groups. All epileptic rats were video-monitored to assess their number of seizures. Epileptic rats with an important seizure reduction (>50%) were classified as responders. SV2A, vesicular γ-aminobutyric acid transporter and vesicular glutamate transporter (VGLUT) expression were assessed by immunostaining. SV2A expression was not modified during epilepsy. However, responders showed ≈55% SV2A-VGLUT co-expression in comparison with the non-responder group (≈40%). Thus, SV2A expression in glutamatergic terminals may be important for the response to LEV treatment.

Behavioral role of PACAP signaling reflects its selective distribution in glutamatergic and GABAergic neuronal subpopulations.

The neuropeptide PACAP, acting as a co-transmitter, increases neuronal excitability, which may enhance anxiety and arousal associated with threat conveyed by multiple sensory modalities. The distribution of neurons expressing PACAP and its receptor, PAC1, throughout the mouse nervous system was determined, in register with expression of glutamatergic and GABAergic neuronal markers, to develop a coherent chemoanatomical picture of PACAP role in brain motor responses to sensory input. A circuit role for PACAP was tested by observing Fos activation of brain neurons after olfactory threat cue in wild-type and PACAP knockout mice. Neuronal activation and behavioral response, were blunted in PACAP knock-out mice, accompanied by sharply downregulated vesicular transporter expression in both GABAergic and glutamatergic neurons expressing PACAP and its receptor. This report signals a new perspective on the role of neuropeptide signaling in supporting excitatory and inhibitory neurotransmission in the nervous system within functionally coherent polysynaptic circuits.

VGLUT-VGAT expression delineates functionally specialised populations of vasopressin-containing neurones including a glutamatergic perforant path-projecting cell group to the hippocampus in rat and mouse brain.

The origin and functional significance of vasopressin (AVP)-containing fibres in limbic regions has been an ongoing subject of investigation for several years. We have previously identified AVP-magnocellular neurones of rat hypothalamus that provide glutamatergic projections to the hippocampus, amygdala, lateral habenula and locus coeruleus. However, we also reported AVP-immunopositive fibres in those regions that are thin and make Gray type II synapses, which are unlikely to be of magnocellular origin. Therefore, in the present study, we characterised AVP mRNA co-expression with expression of mRNAs marking glutamatergic (vesicular glutamate transporter [VGLUT]) and GABAergic (vesicular GABA transporter [VGAT]) neuronal traits in rat and mouse brain, using high-resolution in situ hybridisation methods, including a radio-ribonucleotide and RNAscope 2.5 HD duplex assay, with Slc17a7, Slc17a6, Slc32a1 and Avp probes corresponding to mRNAs of VGLUT1, VGLUT2, VGAT and AVP, respectively. We located 18 cell groups expressing Avp and identified their molecular signatures for VGLUT and VGAT mRNA expression. Avp cell groups of hypothalamus and midbrain are mainly VGLUT mRNA-expressing, whereas those in regions derived from cerebral nuclei are mainly VGAT mRNA-expressing, suggesting a functional segregation of glutamate/GABA co-transmission with AVP. A newly identified Slc17a7 and Slc17a6 (but not Slc32a1) expressing vasopressinergic cell group was found in layer II-III neurones of the central entorhinal cortex, which projects to the hippocampus. These data support the notion of a complex role for AVP with respect to modulating multiple central circuits controlling behaviour in specific ways depending on co-transmission with glutamate or GABA, potentially giving rise to a functional classification of AVPergic neurones in the central nervous system.

Excitatory and inhibitory innervation of the mouse orofacial motor nuclei: A stereological study.

Neurons in the trigeminal (Mo5), facial (Mo7), ambiguus (Amb), and hypoglossal (Mo12) motor nuclei innervate jaw, facial, pharynx/larynx/esophagus, and tongue muscles, respectively. They are essential for movements subserving feeding, exploration of the environment, and social communication. These neurons are largely controlled by sensory afferents and premotor neurons of the reticular formation, where central pattern generator circuits controlling orofacial movements are located. To provide a description of the orofacial nuclei of the adult mouse and to ascertain the influence of excitatory and inhibitory afferents upon them, we used stereology to estimate the number of motoneurons as well as of varicosities immunopositive for glutamate (VGluT1+, VGluT2+) and GABA/glycine (known as VIAAT+ or VGAT+) vesicular transporters in the Mo5, Mo7, Amb, and Mo12. Mo5, Mo7, Amb, and Mo12 contain ∼1,000, ∼3,000, ∼600, and ∼1,700 cells, respectively. VGluT1+, VGluT2+, and VIAAT+ varicosities respectively represent: 28%, 41%, and 31% in Mo5; 2%, 49%, and 49% in Mo7; 12%, 42%, and 46% in Amb; and 4%, 54%, and 42% in Mo12. The Mo5 jaw-closing subdivision shows the highest VGluT1+ innervation. Noticeably, the VGluT2+ and VIAAT+ varicosity density in Mo7 is 5-fold higher than in Mo5 and 10-fold higher than in Amb and Mo12. The high density of terminals in Mo7 likely reflects the convergence and integration of numerous inputs to motoneurons subserving the wide range of complex behaviors to which this nucleus contributes. Also, somatic versus neuropil location of varicosities suggests that most of these afferents are integrated in the dendritic trees of Mo7 neurons.