Probes for INS

Glutamatergic neurons that express pre-proglucagon (PPG) and are immunopositive (+) for glucagon-like peptide-1 (i.e., GLP-1+ neurons) are located within the caudal nucleus of the solitary tract (cNTS) and medullary reticular formation in rats and mice. GLP-1 neurons give rise to an extensive central network in which GLP-1 receptor (R) signaling suppresses food intake, attenuates rewarding, increases avoidance, and stimulates stress responses, partly via . GLP-1R signaling within the cNTS. In mice, noradrenergic (A2) cNTS neurons express GLP-1R, whereas PPG neurons do not. In the present study, confocal microscopy in rats confirmed that prolactin-releasing peptide (PrRP)+ A2 neurons are closely apposed by GLP-1+ axonal varicosities. Surprisingly, GLP-1+ appositions were also observed on dendrites of PPG/GLP-1+ neurons in both species, and electron microscopy in rats revealed that GLP-1+ boutons form asymmetric synaptic contacts with GLP-1+ dendrites. However, RNAscope confirmed that rat GLP-1 neurons do not express GLP-1R mRNA. Similarly, Ca2+ imaging of somatic and dendritic responses in mouse ex vivo slices confirmed that PPG neurons do not respond directly to GLP-1, and a mouse cross-breeding strategy revealed that fewer than 1% of PPG neurons co-express GLP-1R. Collectively, these data suggest that GLP-1R signaling pathways modulate the activity of PrRP+ A2 neurons, and also reveal a local "feed-forward" synaptic network among GLP-1 neurons that apparently does not utilize GLP-1R signaling. This local GLP-1 network may instead use glutamatergic signaling to facilitate dynamic and potentially selective recruitment of GLP-1 neural populations that shape behavioral and physiological responses to internal and external challenges.

Place cells in the CA1 region of the hippocampus express location-specific firing despite receiving a steady barrage of heterogeneously tuned excitatory inputs that should compromise output dynamic range and timing. We examined the role of synaptic inhibition in countering the deleterious effects of off-target excitation. Intracellular recordings in behaving mice demonstrate that bimodal excitation drives place cells, while unimodal excitation drives weaker or no spatial tuning in interneurons. Optogenetic hyperpolarization of interneurons had spatially uniform effects on place cell membrane potential dynamics, substantially reducing spatial selectivity. These data and a computational model suggest that spatially uniform inhibitory conductance enhances rate coding in place cells by suppressing out-of-field excitation and by limiting dendritic amplification. Similarly, we observed that inhibitory suppression of phasic noise generated by out-of-field excitation enhances temporal coding by expanding the range of theta phase precession. Thus, spatially uniform inhibition allows proficient and flexible coding in hippocampal CA1 by suppressing heterogeneously tuned excitation.

Activity-dependent transcriptional responses shape cortical function. However, a comprehensive understanding of the diversity of these responses across the full range of cortical cell types, and how these changes contribute to neuronal plasticity and disease, is lacking. To investigate the breadth of transcriptional changes that occur across cell types in the mouse visual cortex after exposure to light, we applied high-throughput single-cell RNA sequencing. We identified significant and divergent transcriptional responses to stimulation in each of the 30 cell types characterized, thus revealing 611 stimulus-responsive genes. Excitatory pyramidal neurons exhibited inter- and intralaminar heterogeneity in the induction of stimulus-responsive genes. Non-neuronal cells showed clear transcriptional responses that may regulate experience-dependent changes in neurovascular coupling and myelination. Together, these results reveal the dynamic landscape of the stimulus-dependent transcriptional changes occurring across cell types in the visual cortex; these changes are probably critical for cortical function and may be sites of deregulation in developmental brain disorders.

Eating is a learned process. Our desires for specific foods arise through experience. Both electrical stimulation and optogenetic studies have shown that increased activity in the lateral hypothalamus (LH) promotes feeding. Current dogma is that these effects reflect a role for LH neurons in the control of the core motivation to feed, and their activity comes under control of forebrain regions to elicit learned food-motivated behaviors. However, these effects could also reflect the storage of associative information about the cues leading to food in LH itself. Here, we present data from several studies that are consistent with a role for LH in learning. In the first experiment, we use a novel GAD-Cre rat to show that optogenetic inhibition of LH γ-aminobutyric acid (GABA) neurons restricted to cue presentation disrupts the rats' ability to learn that a cue predicts food without affecting subsequent food consumption. In the second experiment, we show that this manipulation also disrupts the ability of a cue to promote food seeking after learning. Finally, we show that inhibition of the terminals of the LH GABA neurons in ventral-tegmental area (VTA) facilitates learning about reward-paired cues. These results suggest that the LH GABA neurons are critical for storing and later disseminating information about reward-predictive cues.

Primary carcinomas of the urethra are rare and poorly understood lesions, hence their clinical and pathologic spectrum is not completely defined. We analyzed a series of 130 primary urethral tumors and classified 106 of them as primary urethral carcinomas. The age at diagnosis of patients with primary urethral carcinomas ranged from 42-97years (mean: 69.4yrs.; median: 70yrs). There were 73 males and 33 female patients with a ratio of 2.2:1. In male patients the tumors most frequently developed in the bulbous-membranous segment of the urethra. In female patients the entire length of the urethra was typically involved. Microscopically, they were poorly differentiated carcinoma with hybrid squamous and urothelial features and developed from precursor intraepithelial conditions such as dysplasia and carcinoma in situ, which were frequently present in the adjacent urethral mucosa. High risk HPV infection could be documented in 31.6% of these tumors. Follow-up information was available for 95 patients. Twenty-three patients died of the disease with a mean and median survival of 39 and 21months respectively. Urethral carcinomas are aggressive tumors with high propensity for regional and distant metastases with mean and median survival of 39 and 21months respectively. Our observations have important implications for the management of patients with primary carcinoma of the urethra by defining them as a unique entity linked to HPV infection.

The dentate gyrus of the hippocampus is a brain region in which neurogenesis persists into adulthood; however, the relationship between developmental and adult dentate gyrus neurogenesis has not been examined in detail. Here we used single-cell RNA sequencing to reveal the molecular dynamics and diversity of dentate gyrus cell types in perinatal, juvenile, and adult mice. We found distinct quiescent and proliferating progenitor cell types, linked by transient intermediate states to neuroblast stages and fully mature granule cells. We observed shifts in the molecular identity of quiescent and proliferating radial glia and granule cells during the postnatal period that were then maintained through adult stages. In contrast, intermediate progenitor cells, neuroblasts, and immature granule cells were nearly indistinguishable at all ages. These findings demonstrate the fundamental similarity of postnatal and adult neurogenesis in the hippocampus and pinpoint the early postnatal transformation of radial glia from embryonic progenitors to adult quiescent stem cells.

Current understanding of the contribution of C1 neurons to blood pressure (BP) regulation derives predominantly from experiments carried out in anesthetized animals or reduced ex vivo preparations. Here we use ArchaerhodopsinT3.0 (ArchT) loss-of-function optogenetics to explore BP regulation by C1 neurons in intact unanesthetized rats. Using a lentivirus that expresses ArchT under the Phox2b-activated promoter PRSx8 (PRSx8-ArchT), ∼65% of transduced neurons were C1 (balance retrotrapezoid nucleus, RTN). Other rats received CaMKII-ArchT3.0 AAV2 (CaMKII-ArchT) which transduced C1 neurons and larger numbers of unidentified glutamatergic and GABAergic cells.Under anesthesia, ArchT-photoactivation reduced sympathetic nerve activity and BP and silenced/strongly inhibited most (7/12) putative C1 neurons. In unanesthetized PRSx8-ArchT-treated rats breathing room air, bilateral ArchT-photoactivation caused a very small BP reduction that was only slightly larger under hypercapnia (6% FiCO2) but was greatly enhanced during hypoxia (10 and 12% FiO2), after sino-aortic denervation, or during isoflurane anesthesia. Degree of hypotension correlated with percentage of ArchT-transduced C1 cells. ArchT-photoactivation produced similar BP changes in CaMKII-ArchT-treated rats. Photoactivation in PRSX8-ArchT rats reduced breathing frequency (FR); in CamKII-ArchT rats FR increased.We conclude that the BP drop elicited by ArchT activation resulted from C1 inhibition and was unrelated to breathing changes. C1 neurons have low activity under normoxia but their activation is important to BP stability during hypoxia or anesthesia and contributes greatly to the hypertension caused by baroreceptor deafferentation. Finally, C1 neurons are marginally activated by hypercapnia and the large breathing stimulation caused by this stimulus has very little impact on resting BP.SIGNIFICANCE STATEMENTC1 neurons (C1) are glutamatergic/peptidergic/catecholaminergic neurons located in the medulla oblongata, which may operate as a switchboard for differential, behavior-appropriate, activation of selected sympathetic efferents. Based largely on experimentation in anesthetized or reduced preparations, a rostrally-located subset of C1 may contribute to both BP stabilization and dysregulation (hypertension). Here we used Archaerhodopsin-based loss-of-function optogenetics to explore the contribution of these neurons to BP in conscious rats. The results suggest that C1 contributes little to resting BP under normoxia or hypercapnia, C1 discharge is continuously restrained by arterial baroreceptors and C1 activation is critical to stabilize BP under hypoxia or anesthesia. This optogenetic approach could also be useful to explore the role of C1 during specific behaviors or in hypertensive models.