Probes for INS

Although Netos are considered auxiliary subunits critical for kainate receptor (KAR) function, direct evidence for their regulation of native KARs is limited. Because Neto KAR regulation is GluK subunit/Neto isoform specific, such regulation must be determined in cell-type-specific contexts. We demonstrate Neto1/2 expression in somatostatin (SOM)-, cholecystokinin/cannabinoid receptor 1 (CCK/CB1)-, and parvalbumin (PV)-containing interneurons. KAR-mediated excitation of these interneurons is contingent upon Neto1 because kainate yields comparable effects in Neto2 knockouts and wild-types but fails to excite interneurons or recruit inhibition in Neto1 knockouts. In contrast, presynaptic KARs in CCK/CB1 interneurons are dually regulated by both Neto1 and Neto2. Neto association promotes tonic presynaptic KAR activation, dampening CCK/CB1 interneuron output, and loss of this brake in Neto mutants profoundly increases CCK/CB1 interneuron-mediatedinhibition. Our results confirm that Neto1 regulates endogenous somatodendritic KARs in diverse interneurons and demonstrate Neto regulation of presynaptic KARs in mature inhibitory presynaptic terminals.

The medial septum (MS) complex modulates hippocampal function and related behaviors. Septohippocampal projections promote and control different forms of hippocampal synchronization. Specifically, GABAergic and cholinergic projections targeting the hippocampal formation from the MS provide bursting discharges to promote theta rhythm, or tonic activity to promote gamma oscillations. In turn, the MS is targeted by ascending projections from the hypothalamus and brainstem. One of these projections arises from the nucleus incertus in the pontine tegmentum, which contains GABA neurons that co-express the neuropeptide relaxin-3 (Rln3). Both stimulation of the nucleus incertus and septal infusion of Rln3 receptor agonist peptides promotes hippocampal theta rhythm. The Gi/o-protein-coupled receptor, relaxin-family peptide receptor 3 (RXFP3), is the cognate receptor for Rln3 and identification of the transmitter phenotype of neurons expressing RXFP3 in the septohippocampal system can provide further insights into the role of Rln3 transmission in the promotion of septohippocampal theta rhythm. Therefore, we used RNAscope multiplex in situ hybridization to characterize the septal neurons expressing Rxfp3mRNA in the rat. Our results demonstrate that Rxfp3 mRNA is abundantly expressed in vesicular GABA transporter (vGAT) mRNA- and parvalbumin (PV) mRNA-positive GABA neurons in MS, whereas ChATmRNA-positive acetylcholine neurons lack Rxfp3 mRNA. Approximately 75% of Rxfp3 mRNA-positive neurons expressed vGAT mRNA (and 22% were PV mRNA-positive), while the remaining 25% expressed Rxfp3 mRNA only, consistent with a potential glutamatergic phenotype. Similar proportions were observed in the posterior septum. The occurrence of RXFP3 in PV-positive GABAergic neurons gives support to a role for the Rln3-RXFP3 system in septohippocampal theta rhythm.

The histone H3 lysine 27 (H3K27) demethylase Kdm6b (Jmjd3) can promote cellular differentiation, however its physiological functions in neurons remain to be fully determined. We studied the expression and function of Kdm6b in differentiating granule neurons of the developing postnatal mouse cerebellum. At postnatal day 7, Kdm6b is expressed throughout the layers of the developing cerebellar cortex, but its expression is upregulated in newborn cerebellar granule neurons (CGNs). Atoh1-Cre mediated conditional knockout of Kdm6b in CGN precursors either alone or in combination with Kdm6a did not disturb the gross morphological development of the cerebellum. Furthermore, RNAi-mediated knockdown of Kdm6b in cultured CGN precursors did not alter the induced expression of early neuronal marker genes upon cell cycle exit. By contrast, knockdown of Kdm6b significantly impaired the induction of a mature neuronal gene expression program, which includes gene products required for functional synapse maturation. Loss of Kdm6b also impaired the ability of Brain-Derived Neurotrophic Factor (BDNF) to induce expression of Grin2c and Tiam1 in maturing CGNs. Taken together, these data reveal a previously unknown role for Kdm6b in the postmitotic stages of CGN maturation and suggest that Kdm6b may work, at least in part, by a transcriptional mechanism that promotes gene sensitivity to regulation by BDNF.

Proprioception, the perception of body and limb position, is mediated by proprioceptors, specialized mechanosensory neurons that convey information about the stretch and tension experienced by muscles, tendons, skin and joints. In mammals, the molecular identity of the stretch-sensitive channel that mediates proprioception is unknown. We found that the mechanically activated nonselective cation channel Piezo2 was expressed in sensory endings of proprioceptors innervating muscle spindles and Golgi tendon organs in mice. Two independent mouse lines that lack Piezo2 in proprioceptive neurons showed severely uncoordinated body movements and abnormal limb positions. Moreover, the mechanosensitivity of parvalbumin-expressing neurons that predominantly mark proprioceptors was dependent on Piezo2 expression in vitro, and the stretch-induced firing of proprioceptors in muscle-nerve recordings was markedly reduced in Piezo2-deficient mice. Together, our results indicate that Piezo2 is the major mechanotransducer of mammalian proprioceptors.

Oxytocin receptor (Oxtr) signaling in neural circuits mediating discrimination of social stimuli and affiliation or avoidance behavior is thought to guide social recognition. Remarkably, the physiological functions of Oxtrs in the hippocampus are not known. Here we demonstrate using genetic and pharmacological approaches that Oxtrs in the anterior dentate gyrus (aDG) and anterior CA2/CA3 (aCA2/CA3) of mice are necessary for discrimination of social, but not non-social, stimuli. Further, Oxtrs in aCA2/CA3 neurons recruit a population-based coding mechanism to mediate social stimuli discrimination. Optogenetic terminal-specific attenuation revealed a critical role for aCA2/CA3 outputs to posterior CA1 for discrimination of social stimuli. In contrast, aCA2/CA3 projections to aCA1 mediate discrimination of non-social stimuli. These studies identify a role for an aDG-CA2/CA3 axis of Oxtr expressing cells in discrimination of social stimuli and delineate a pathway relaying social memory computations in the anterior hippocampus to the posterior hippocampus to guide social recognition.

Major depressive disorder (MDD) patients display a common but often variable set of symptoms making successful, sustained treatment difficult to achieve. Separate depressive symptoms may be encoded by differential changes in distinct circuits in the brain, yet how discrete circuits underlie behavioral subsets of depression and how they adapt in response to stress has not been addressed. We identify two discrete circuits of parvalbumin-positive (PV) neurons in the ventral pallidum (VP) projecting to either the lateral habenula or ventral tegmental area contributing to depression. We find that these populations undergo different electrophysiological adaptations in response to social defeat stress, which are normalized by antidepressant treatment. Furthermore, manipulation of each population mediates either social withdrawal or behavioral despair, but not both. We propose that distinct components of the VP PV circuit can subserve related, yet separate depressive-like phenotypes in mice, which could ultimately provide a platform for symptom-specific treatments of depression.

In human prefrontal cortex (PFC), ~85% of γ-aminobutyric acid (GABA)-expressing neurons can be subdivided into non-overlapping groups by the presence of calbindin (CB), calretinin (CR) or parvalbumin (PV). Substantial research has focused on the differences in the laminar locations of the cells bodies of these neurons, with limited attention to the distribution of their axon terminals, their sites of action. We previously reported that in non-human primates subtypes of these cells are distinguishable by differences in terminal protein levels of the GABA synthesizing enzymes glutamic acid decarboxylase 65 (GAD65) and GAD67. Here we used multi-label fluorescence microscopy in human PFC to assess: (1) the laminar distributions of axon terminals containing CB, CR, or PV; and (2) the relative protein levels of GAD65, GAD67 and vesicular GABA transporter (vGAT) in CB, CR and PV terminals. The densities of the different CB, CR and PV terminal subpopulations differed across layers of the PFC. PV terminals comprised two subsets based on the presence of only GAD67 (GAD67+) or both GADs (GAD65/GAD67+), whereas CB and CR terminals comprised three subsets (GAD65+, GAD67+, or GAD65/GAD67+). The densities of the different CB, CR and PV GAD terminal subpopulations also differed across layers. Finally, within each of the three calcium-binding protein subpopulations intra-terminal protein levels of GAD and vGAT differed by GAD subpopulation. These findings are discussed in the context of the laminar distributions of CB, CR and PV cell bodies and the synaptic targets of their axons.