Probes for INS

Cocaine and Amphetamine-regulated Transcript (CART) is one of the most abundant neuropeptides in vagal afferents, including those involved in regulating feeding. Recent observations indicate that metabolic challenges dramatically alter the neuropeptidergic profile of CART-producing vagal afferents. Here, using confocal microscopy, we re-assessed the distribution and regulation of CART (55-102) immunoreactivity in vagal afferents of the male mouse in response to metabolic challenges, including fasting, high-fat diet feeding. Importantly, the perikarya and axons of vagal C-fibers were labeled using mice expressing channelrodhopsin-2 (ChR2-YFP) in Nav1.8-Cre-expressing neurons. In these mice, approximately 82% of the nodose ganglion neurons were labeled with ChR2-YFP. Furthermore, ChR2-YFP-labeled axons could easily be identified in the dorsovagal complex. CART (55-102) immunoreactivity was observed in 55% of the ChR2-YFP-labeled neurons in the nodose ganglion and 22% of the ChR2-YFP-labeled varicosities within the area postrema of fed, fasted and obese mice. The distribution of positive profiles was also identical across the full range of CART staining in fed, fasted and obese mice. In contrast to previous studies, fasting did not induce melanin-concentrating hormone immunoreactivity in vagal afferents. Moreover, prepro-MCH mRNA was undetectable in the nodose ganglion of fasted mice. In summary, this study showed that the perikarya and central terminals of vagal afferents are invariably enriched in CART and devoid of MCH.

Significance Statement Recent studies reported that fasting triggers vagal afferents to switch from expressing anorectic to orexigenic neuropeptides. This study failed to replicate the aforementioned observations using a combination of confocal microscopy, immunohistochemistry, and in situ hybridization. In particular, we showed that neither fasting nor diet-induced obesity influence the immunoreactivity for Cocaine and Amphetamine-regulated Transcript neuropeptide in the mouse vagal afferents. In contrast to previous studies, we also failed to detect melanin-concentrating hormone expression in the mouse vagal afferents. Overall, we reached the conclusion that the neuropeptidergic profile of the vagal afferents involved in feeding is remarkably stable in response to metabolic challenges.

In the brain, purines such as ATP and adenosine can function as neurotransmitters and co-transmitters, or serve as signals in neuron-glial interactions. In thalamocortical (TC) projections to sensory cortex, adenosine functions as a negative regulator of glutamate release via activation of the presynaptic adenosine A1 receptor (A1 R). In the auditory forebrain, restriction of A1 R-adenosine signaling in medial geniculate (MG) neurons is sufficient to extend LTP, LTD, and tonotopic map plasticity in adult mice for months beyond the critical period. Interfering with adenosine signaling in primary auditory cortex (A1) does not contribute to these forms of plasticity, suggesting regional differences in the roles of A1 R-mediated adenosine signaling in the forebrain. To advance understanding of the circuitry, in situ hybridization was used to localize neuronal and glial cell types in the auditory forebrain that express A1 R transcripts (Adora1), based on co-expression with cell-specific markers for neuronal and glial subtypes. In A1, Adora1 transcripts were concentrated in L3/4 and L6 of glutamatergic neurons. Subpopulations of GABAergic neurons, astrocytes, oligodendrocytes, and microglia expressed lower levels of Adora1. In MG, Adora1 was expressed by glutamatergic neurons in all divisions, and subpopulations of all glial classes. The collective findings imply that A1 R-mediated signaling broadly extends to all subdivisions of auditory cortex and MG. Selective expression by neuronal and glial subpopulations suggests that experimental manipulations of A1 R-adenosine signaling could impact several cell types, depending on their location. Strategies to target Adora1 in specific cell types can be developed from the data generated here.

Numerous genetic and functional studies implicate variants of Neuregulin-1 (NRG1) and its neuronal receptor ErbB4 in schizophrenia and many of its endophenotypes. Although the neurophysiological and behavioral phenotypes of NRG1 mutant mice have been investigated extensively, practically nothing is known about the function of NRG2, the closest NRG1 homolog. We found that NRG2 expression in the adult rodent brain does not overlap with NRG1 and is more extensive than originally reported, including expression in the striatum and medial prefrontal cortex (mPFC), and therefore generated NRG2 knockout mice (KO) to study its function. NRG2 KOs have higher extracellular dopamine levels in the dorsal striatum but lower levels in the mPFC; a pattern with similarities to dopamine dysbalance in schizophrenia. Like ErbB4 KO mice, NRG2 KOs performed abnormally in a battery of behavioral tasks relevant to psychiatric disorders. NRG2 KOs exhibit hyperactivity in a novelty-induced open field, deficits in prepulse inhibition, hypersensitivity to amphetamine, antisocial behaviors, reduced anxiety-like behavior in the elevated plus maze and deficits in the T-maze alteration reward test-a task dependent on hippocampal and mPFC function. Acute administration of clozapine rapidly increased extracellular dopamine levels in the mPFC and improved alternation T-maze performance. Similar to mice treated chronically with N-methyl-d-aspartate receptor (NMDAR) antagonists, we demonstrate that NMDAR synaptic currents in NRG2 KOs are augmented at hippocampal glutamatergic synapses and are more sensitive to ifenprodil, indicating an increased contribution of GluN2B-containing NMDARs. Our findings reveal a novel role for NRG2 in the modulation of behaviors with relevance to psychiatric disorders.

Stressful life events induce abnormalities in emotional and cognitive behaviour. The endogenous opioid system plays an essential role in stress adaptation and coping strategies. In particular, the µ-opioid receptor (μR), one of the major opioid receptors, strongly influences memory processing in that alterations in μR signalling are associated with various neuropsychiatric disorders. However, it remains unclear whether μR signalling contributes to memory impairments induced by acute stress. Here, we utilized pharmacological methods and cell-type-selective/non-cell-type-selective μR depletion approaches combined with behavioural tests, biochemical analyses, and in vitro electrophysiological recordings to investigate the role of hippocampal μR signalling in memory-retrieval impairment induced by acute elevated platform (EP) stress in mice. Biochemical and molecular analyses revealed that hippocampal μRs were significantly activated during acute stress. Blockage of hippocampal μRs, non-selective deletion of μRs or selective deletion of μRs on GABAergic neurons (μRGABA) reversed EP-stress-induced impairment of memory retrieval, with no effect on the elevation of serum corticosterone after stress. Electrophysiological results demonstrated that stress depressed hippocampal GABAergic synaptic transmission to CA1 pyramidal neurons, thereby leading to excitation/inhibition (E/I) imbalance in a μRGABA-dependent manner. Pharmaceutically enhancing hippocampal GABAAreceptor-mediated inhibitory currents in stressed mice restored their memory retrieval, whereas inhibiting those currents in the unstressed mice mimicked the stress-induced impairment of memory retrieval. Our findings reveal a novel pathway in which endogenous opioids recruited by acute stress predominantly activate μRGABA to depress GABAergic inhibitory effects on CA1 pyramidal neurons, which subsequently alters the E/I balance in the hippocampus and results in impairment of memory retrieval.

Social recognition, the ability to recognize individuals that were previously encountered, requires complex integration of sensory inputs with previous experience. Here, we use a variety of approaches to discern how oxytocin sensitive neurons in the prefrontal cortex (PFC) exert descending control over a circuit mediating social recognition in mice. Using male mice with Cre-recombinase directed to the oxytocin receptor gene (Oxtr), we revealed that the Oxtr is expressed on glutamatergic neurons in the PFC, optogenetic stimulation of which, elicited activation of neurons residing in several mesolimbic brain structures. Optogenetic stimulation of axons in the basolateral amygdala (BLA) arising from Oxtr-expressing neurons in the PFC eliminated the ability to distinguish novel from familiar conspecifics, but remarkably, distinguishing between novel and familiar objects was unaffected. These results suggest that an oxytocin sensitive PFC to BLA circuit is required for social recognition. The implication is that impaired social memory may manifest from dysregulation of this circuit.SIGNIFICANCE STATEMENTUsing mice we demonstrate that optogenetic activation of the neurons in the prefrontal cortex (PFC) that express the oxytocin receptor gene (Oxtr) impairs the ability to distinguish between novel and familiar conspecifics but the ability to distinguish between novel and familiar objects remains intact. Subjects with Autism Spectrum Disorders (ASD) have difficulty identifying a person based on remembering facial features; however, ASD and typical subjects perform similarly when remembering objects. In subjects with ASD, viewing the same face increases neural activity in the PFC, which may be analogous to the optogenetic excitation of Oxtr-expressing neurons in the PFC that impairs social recognition in mice. The implication is that over-activation of Oxtr-expressing neurons in the PFC may contribute to ASD symptomology.

The goals of this study were to localize the neuropeptide Cocaine- and Amphetamine-Regulated Transcript (CART) in primate retinas and to describe the morphology, neurotransmitter content and synaptic connections of the neurons that contain it. Using in situ hybridization, light and electron microscopic immunolabeling, CART was localized to GABAergic amacrine cells in baboon retinas. The CART-positive cells had thin, varicose dendrites that gradually descended through the inner plexiform layer and ramified extensively in the innermost stratum. They resembled two types of wide-field diffuse amacrine cells that had been described previously in macaque retinas using the Golgi method and also A17, serotonin-accumulating and waterfall cells of other mammals. The CART-positive cells received synapses from rod bipolar cell axons and made synapses onto the axons in a reciprocal configuration. The CART-positive cells also received synapses from other amacrine cells. Some of these were located on their primary dendrites, and the presynaptic cells there included dopaminergic amacrine cells. Although some CART-positive somas were localized in the ganglion cell layer, they did not contain the ganglion cell marker RNA binding protein with multiple splicing (RBPMS). Based on these results and electrophysiological studies in other mammals, the CART-positive amacrine cells would be expected to play a major role in the primary rod pathway of primates, providing feedback inhibition to rod bipolar cells.