Alcohol withdrawal (AW) after chronic alcohol exposure (AE) produces a series of symptoms. Among them,
generalized tonic-clonic seizures are the most severe and dangerous symptom. The severity and susceptibility
to relapse, and perpetuation of alcohol abuse underscore the urgent need to understand mechanisms underlying
alcohol dependence and AW in order to develop new therapeutic strategies to intervene and treat AW-associated
syndromes such as seizures. In this application, we will test the novel hypothesis that activity and connectivity
of hippocampal newborn dentate granule cells (DGCs) underlie AW-associated seizures. DGCs are principal
excitatory neurons that are continuously produced and integrate into hippocampal neural circuits, and altered
hippocampal neurogenesis has been implicated in seizures. Our previous studies have revealed the essential
roles of hippocampal newborn DGCs in the expressions of AW-associated seizures. AE reduced spine formation
while AW increased synaptic connectivity of hippocampal newborn DGCs. Our rabies virus-mediated retrograde
tracing study discovered altered neuronal connectivity of hippocampal newborn DGCs with both excitatory and
inhibitory neurons during AW seizures. Moreover, our functional study with a DREADD (Designer Receptors
Exclusively Activated by Designer Drugs) method demonstrated that activity of hippocampal newborn neurons
plays an essential role in the expression of AW-associated seizures. These observations provided the theoretical
foundation for our hypothesis that altered neuronal connectivity and activity of hippocampal newborn DGCs
disrupts the balance of excitatory and inhibitory (E/I) signals, ultimately leading to AW-associated seizures. Thus,
the central goal of this proposal is to use novel mapping methods, imaging tools, and cellular and molecular
approaches in order to understand activity and connectivity of hippocampal neural circuits that are responsible
for AW-associated seizures. In Aim 1, we will determine whether AW alters neuronal and functional connectivity
of DGCs by using a rabies virus- and wheat germ agglutinin (WGA)-mediated retrograde and anterograde tracing
methods, respectively. We will also use multiple DREADDs and assess the essential role of de novo neural
circuits formed between hippocampal newborn DGCs and input neurons in AW-associated seizures. Aim 2, using
Ca2+ imaging and various magnetic resonance imaging (MRI) modalities that allows us for longitudinal studies,
we will determine activity and connectivity of hippocampal and global neural circuits underlying AW-associated
seizures. In Aim 3, we will identify and validate transcriptome that may underlie altered synaptic and neuronal
connectivity of hippocampal newborn DGCs in response to AE and AW. The usage of single cell RNA sequencing
will allow us to not only register differentially expressed genes to cell type-specific manner, but determine
transcriptomes that distinguish pathological newborn DGCs from normal DGCs during AE and AW. Altogether,
our proposal will dissect the molecular, cellular, and neural circuitry mechanisms by which hippocampal newborn
DGCs underlie AW-associated seizures.
