Molecular signaling during development and maturation of the nervous system

National Institute Of Neurological Disorders And Stroke

Investigators

Linked publications, trials & patents

Abstract

Objective 1: define the neuronal Nrg1 signaling network. Subobjective 1: Axonal isoforms of Nrg1 participate in both local axonal signaling and in axon-to-nucleus signaling. To identify the suite of genes targeted by Nrg1 nuclear signaling, we generated RNAseq data from dentate gyrus, nucleus accumbens, basal lateral amygdala, total hippocampus and frontal cortices of mice carrying a mutation in Nrg1 that impairs nuclear signaling (initially identified as a psychosis risk gene in a Costa Rican population). Our most in depth analyses focused on the young adult dentate gyrus; a brain region that undergoes neurogenesis throughout life, allowing us to examine effects of disrupted Nrg1 nuclear signaling in neuroprogenitor populations as well as in mature neurons. Three major findings of these studies are: A) The population of genes whose expression is significantly altered in the absence of Nrg1 nuclear signaling overlaps significantly with populations of genes whose altered expression is associated with risk of being diagnosed with psychotic illness (collaboration with Dr. Kory Johnson, NINDS bioinformatics core; Rajebhosale et al., BioRxiv 2022). B) Loss of Nrg1 nuclear signaling alters the fate specification of neurons in the dentate gyrus resulting in overproduction and ectopic location of the normally rare semi-lunar granule cell population. Strikingly, this phenotype resembles reported effects seen in other mouse models of psychiatric illness, both genetic and environmental. C) The suite of genes that are differentially expressed in mice lacking Nrg1 nuclear signaling shows little overlap with those seen in animals that are heterozygous for genetic disruption of axonal Nrg1, despite the latter animals displaying marked synaptic, circuit and behavioral abnormalities --- a finding implicating a major role for local axonal Nrg1 signaling that is more sensitive to gene dosage than Nrg1 axon to nucleus signaling. Subobjective 2: Prior findings implicate local axonal Nrg1 signaling in axon outgrowth, synapse formation and formation of functional pre-synaptic neurotransmitter release sites (glutamatergic). Ongoing studies delve further into the molecular mechanisms underlying these deficits. Results during the past year demonstrate that local axonal Nrg1 signaling regulates mitochondria trafficking and fission-fusion dynamics and, surprisingly axonal protein synthesis both processes that are critical for maturation of axonal presynaptic specializations. Objective 2: identify the functional heterogeneity of cholinergic neurons. Subobjective 1: Determine the involvement of cholinergic neurons in distinct behaviors. We have identified specific populations of cholinergic neurons that participate in distinct types of memory and demonstrated for the first time that cholinergic neurons participate directly in memory engrams (Crouse et al. ELife 2020 and Rajebhosale et al., bioRxiv 2021, under Re-review ELife)(carried out in collaboration with Dr. L Role, NINDS IRP and Dr. M Picciotto, Yale Univ). These findings provide significantly new insight into the ways that the modulatory, cholinergic system participates in memory formation and retrieval with clear implications to understanding age related cognitive decline that accompanies degeneration of cholinergic brain nuclei. We have initiated a line of investigation focusing on the response of ventral pallidal cholinergic neurons to either innately appetitive or innately aversive olfactory stimuli. Both stimuli broadly activate cholinergic neurons in the ventral pallidum. Using intersectional genetic approaches, we have found that although both stimuli activate cholinergic neurons, the cholinergic neurons activated, represent two distinct, non-overlapping populations implicating these cholinergic neurons in encoding valence and not just salience of sensory stimuli. One is activated by innately appetitive stimuli and is essential for the resulting approach behavior, whereas the second is activated by aversive stimuli. This second population is not required for active avoidance behavior but appears to work in concert with other basal forebrain cholinergic neurons to shape response to threat sculpting the choice between fleeing and freezing. We are now embarking on efforts to understand, at the molecular level, the fundamental properties of these distinct populations. Subobjective 2: 2.1. Dissect the molecular underpinnings of BFCN heterogeneity. A collaborative project with Drs. L Role (NINDS), K. Johnson (NINDS) and T Petros (NICHD), has generated a single nucleus RNAseq dataset from 20,000 forebrain cholinergic neurons. This far surpasses any existing data set in probing cholinergic transcriptomes, expanding the number of subpopulations from 2-3 to over a dozen. We are using spatial transcriptomics to anatomically map these populations and are using these data to probe questions related to the maturation of cholinergic circuits and their age-related vulnerabilities that lead to cognitive impairments. 2.2. Understand the contribution of axonal Nrg1 signaling to maturation and maintenance of the basal forebrain cholinergic system. Using our functional and molecular definitions of cholinergic neuron subtypes as guides, we are embarking on a quest to gain a detailed understanding of their generation and maturation and in particular to define the influence of axonal Nrg1 signaling in these processes. Both snRNAseq and spatial transcriptomics demonstrate differential expression of Nrg1 across the basal forebrain. We are using intersectional genetic approaches to determine whether changing Nrg1 signaling alters maturation of different populations of cholinergic neurons, their participation in synaptic modulation in behaviorally relevant circuits and their resilience vs vulnerability to aging. We have also developed a preparation that allows us to study axonal Nrg1 signaling in cholinergic neurons at high resolution in vitro. Early results indicate that reduced Nrg1 signaling differentially alters the maturation and maintenance of distinct subpopulations of these neurons.