Cellular disturbances in the inhibitory and excitatory balance of brain circuits have been implicated in various neurodegenerative disorders such as Alzheimer's Disease (AD), Multiple Sclerosis (MS) and Parkinson's Disease (PD) (Lauterborn et al. 2021; Bi et al. 2020; Vico Varela et al. 2019; Lerdkrai et al. 2018; Campanelli et al. 2022; Ellvardt et al. 2018). Additionally, these imbalances, such as neuronal hyperactivity or hypoactivity, have been demonstrated to be core features of psychiatric disorders, including Post-Traumatic Stress Disorder (PTSD), Schizophrenia, and Major Depressive Disorder (MDD) (Badura-Brack et al. 2018; Fang et al. 2018; Clancy et al. 2017; Heckers & Konradi et al. 2014; Vadodaria et al. 2019; Helm et al. 2018). Interestingly, psychiatric disorders are often comorbid with neurodegenerative diseases, especially in AD and MDD (Martin-Sanchez et al. 2021). Along similar lines, psychiatric disorders such as MDD and PTSD significantly elevate the risk of developing neurodegenerative diseases such as AD (Flatt et al. 2018; Ownby et al, 2006; Byers & Yaffe, 2011). Thus, imbalances in circuit-level activity provide a putative shared mechanism underlying different brain and may serve as a therapeutic target.

Recently, many studies have investigated the role of direct brain modulation for the treatment and prevention of disease (Johnson et al., 2013; Gauthier et al. 2022), though the identity of the perturbed cells remains relatively unknown. For instance, stimulation of brain cells via transmagnetic stimulation (TMS), which is an approved therapy for treatment-resistant MDD, has promising therapeutic value when applied to AD, PD, stroke, schizophrenia, and MS (Weiler et al. 2020; Brys et al. 2016; van Lieshout et al. 2020; Cole et al. 2015; Nasios et al. 2018). However, understanding how to restore brain patterns through a cell-type specific and targeted manner remains elusive, especially when used for the treatment of brain diseases. Hyperactivity is a hallmark of many disorders and often occurs in areas of the brain involved in learning and memory. For instance, the hippocampus (HPC) is a deep brain structure crucially involved in learning and memory, and its connecting sub-regions are crucial for the formation and retrieval of episodic memories in particular (Eichenbaum, Otto & Cohen, 1992; Moser & Moser, 1998; Burgess, Maguire & O'Keefe, 2002; Eichenbaum & Fortin, 2003). Various clinical and preclinical studies have shown that in early disease states of AD, the HPC becomes hyperactive and, therefore, is a prime location for network dysfunction and disconnection from other brain regions (Palop et al. 2007; Busche et al. 2008; Busche et al. 2012; Palop & Mucke, 2016; Klink et al 2021; Anastacio, Matosi, & Ooi, 2022; Kuchibhotla et. al., 2009). One of the major subregions of the HPC that is directly affected in such diseases is along the ventral axis, especially ventral CA1 (vCA1; Maruszak & Thuret, 2014). Notably, the ventral HPC (vHPC) has direct connectivity to many downstream regions such as the basolateral amygdala (BLA), nucleus accumbens (NAc), medial prefrontal cortex (mPFC), and the hypothalamus, thus underscoring its role in modulating key aspects of memory and behavior (Fanselow & Dong, 2010; Gergues et. al., 2020; Ciocchi et. al., 2015; Padilla-Coreano et. al., 2016; Jimenez et. al., 2018; Zhou et. al., 2019; Phillips et. al., 2019). These connections have been specifically linked to processing the emotionally valenced components of episodic memories (Fanselow & Dong, 2010). Fittingly, dysfunction in a major hub like the vHPC may lead to deficiencies in numerous cognitive functions including learning, memory, emotional processing, reward-seeking behaviors, and stress responses. Thus, we sought to modulate vCA1 processing by chronically modulating its activity, to test the hypothesis that prolonged network perturbation would lead to cellular and behavioral abnormalities that may be unique to each cell type.

Within the vHPC, a number of studies have focused specifically on neuronal functioning and characterizing the roles of neurons in social, anxiety, goal-directed and fear-related behaviors (Jimenez et. al., 2018, Okuyama et. al., 2016; Jimenez et. al., 2020; Padilla-Coreano et. al., 2016; Ciocchi et. al., 2015; Fanselow & Dong, 2010). Notably, glial cells such as astrocytes, have remained understudied in this ventral axis of the HPC. Astrocytes have primarily been studied as support cells in the brain, aiding in neuronal metabolism, supporting the blood-brain barrier and providing homeostatic regulation of their local environment (Dehouck et. al., 1990; Simard & Nedergard, 2004; Kofuji & Newman, 2004; Paulson & Newman, 1987; Tsacopoulous & Magistretti, 1996; Wallraff et. al., 2006; Rouach et. al., 2008; Figley et. al., 2011). Their active role in synaptic regulation has now been fully recognized, as knowledge of their expression of receptors, transporters and bidirectional communication with neurons has grown (Porter et. al., 1997; Perea & Araque, 2005; Covelo & Araque, 2018; Araque et. al., 2001; Di Castro et. al., 2011; Haydon et. al., 2001; Bezzi & Volterra, 2001). It is now generally thought that astrocytes dynamically respond and modulate circuit activity via calcium-dependent and -independent release of gliotransmitters, such as glutamate, D-serine, and ATP, at the tripartite synapse (Araque et. al., 1999; Perea et. al., 2009; Parpura et. al., 1994; Koizumi et. al., 2005; Fellin et. al., 2004). As astrocytes play a pivotal role in many brain functions, modulation of their structure or activity has a major impact on cognition and behavior (Adamsky et. al., 2018; Kol et. al., 2020; Li et. al., 2020; Mederos et. al., 2021; Martin-Fernandez et. al., 2017; Skucas et. al., 2011). Recent work has shown that astrocytic calcium dysregulation in the cortex may be involved in the early stages of neurodegeneration and influence the onset of neuronal hyperactivity even before amyloid deposition in a mouse model of AD (Shah et. al., 2022). Astrocytes and neurons are heterogenous in their structure and function within and across brain regions, suggesting that perturbation of either cell type may elicit unique behavioral and cellular responses even within the same brain region (Oberheim, Goldman & Nedergaard, 2012; Matyash & Kettenmann, 2010; Buosi et. al., 2017; Zhang & Barres, 2010).

Here, we sought to study the differential effects of chemogenetic activation of the hM3D(Gq) pathway in excitatory neurons (calcium/calmodulin-dependent protein kinase II; CaMKII+) or astrocytes (glial fibrillary acidic protein; GFAP+) in the vHPC, as most studies have identified network hyper- or hypoactivity as a result of disease rather than how onset of this activity may produce pathological brain functioning. In this study, we tested the hypothesis that artificial chronic induction of network hyperactivity in both neuronal and astrocytic groups is sufficient to induce impairments in cognition and behavior, as well as changes in histological markers of cellular stress and inflammation. Our results demonstrate that chronic Gq pathway activation of CaMKII+ neurons or GFAP+ astrocytes in vHPC across multiple time points is sufficient to induce changes in behavior and histological markers. Behaviorally, CaMKII-Gq activation decreased contextual fear acquisition at the 9 month time point and increased fear during extinction at the 3 month time point. GFAP-Gq activation was generally impacted by our manipulation during CFC, recall and extinction at the 6 and 9 month time points. For anxiety-related tasks, CaMKII-Gq activation displayed a combinatory effect with aging to impact behavior. GFAP-Gq activation decreased anxiety-related behaviors at the 3 month time point in the open field, but only showed impacts of aging in the zero maze. For social interaction, there was a decrease in engagement with the social target in the CaMKII-Gq mice at 6 months. For GFAP-Gq mice, we observed only effects of aging on locomotion and social target engagement in the social interaction task. For novel environment exploration, there was an impact of chronic activation and aging on neuronal and astrocytic groups across all time points. Histologically, CaMKII-Gq mice displayed changes in microglial, but not astrocytic cell number in vHPC, while GFAP-Gq mice showed no differences in glial cell number. For morphology, only GFAP-hM3Dq activation impacted microglial characteristics, but not astrocytic, and CaMKII-hM3Dq activation did not affect either cell type. Our study thus provides a mechanistic approach to study how these cell types can be longitudinally engaged to produce behavioral deficits, which provides a putative link to disorders that have similar characteristic network dysfunctions (e.g. hyper- or hypoactivity), while also adding a more direct role for glia in modulating behavior.