Zolpidem Reduces Hippocampal Neuronal Activity in Freely Behaving Mice: A Large Scale Calcium Imaging Study with Miniaturized Fluorescence Microscope

Abstract

Therapeutic drugs for cognitive and psychiatric disorders are often characterized by their molecular mechanism of action. Here we demonstrate a new approach to elucidate drug action on large-scale neuronal activity by tracking somatic calcium dynamics in hundreds of CA1 hippocampal neurons of pharmacologically manipulated behaving mice. We used an adeno-associated viral vector to express the calcium sensor GCaMP3 in CA1 pyramidal cells under control of the CaMKII promoter and a miniaturized microscope to observe cellular dynamics. We visualized these dynamics with and without a systemic administration of Zolpidem, a GABAA agonist that is the most commonly prescribed drug for the treatment of insomnia in the United States. Despite growing concerns about the potential adverse effects of Zolpidem on memory and cognition, it remained unclear whether Zolpidem alters neuronal activity in the hippocampus, a brain area critical for cognition and memory. Zolpidem, when delivered at a dose known to induce and prolong sleep, strongly suppressed CA1 calcium signaling. The rate of calcium transients after Zolpidem administration was significantly lower compared to vehicle treatment. To factor out the contribution of changes in locomotor or physiological conditions following Zolpidem treatment, we compared the cellular activity across comparable epochs matched by locomotor and physiological assessments. This analysis revealed significantly depressive effects of Zolpidem regardless of the animal’s state. Individual hippocampal CA1 pyramidal cells differed in their responses to Zolpidem with the majority (∼65%) significantly decreasing the rate of calcium transients, and a small subset (3%) showing an unexpected and significant increase. By linking molecular mechanisms with the dynamics of neural circuitry and behavioral states, this approach has the potential to contribute substantially to the development of new therapeutics for the treatment of CNS disorders.

Cellular Level Brain Imaging in Behaving Mammals: An Engineering Approach

“…Wireless electrophysiological recordings from multiple animals in parallel are already widely used in behavioral neuropharmacology studies, and a wireless approach to cellular imaging might notably benefit pre-clinical and therapeutic discovery efforts. To date, brain imaging in freely moving mice has been combined with wireless telemetry systems that transmit electroencephalography (EEG) and electromyography (EMG) signals (Berdyyeva et al, 2014). There have also been efforts to improve the throughput of imaging studies in head-restrained rats, by allowing rats to initiate brief periods of voluntary head restraint underneath the objective lens of a conventional two-photon microscope (Scott et al, 2013).…”

“…Initially, the illumination and fluorescence image propagated to and from the mouse via a fiber-optic bundle (Flusberg et al, 2008; Murayama and Larkum, 2009; Murayama et al, 2007, 2009; Soden et al, 2013), but the subsequent approach of integrating all optical components within the head-mounted device has proven notably superior regarding the optical sensitivity, field of view, resolution, mechanical flexibility for the animal, and portability between experimental sites that can be attained (Ghosh et al, 2011). Using a fingertip-sized integrated microscope (Figure 4A) that combines a blue light-emitting diode (LED), a cell phone CMOS camera chip for digital imaging, miniaturized lenses, and a fluorescence filter set within a single compact housing, one-photon imaging of neural Ca 2+ signals in freely behaving mice has become more widespread since the initial experiments (Figures 4B–4F) (Berdyyeva et al, 2014; Ghosh et al, 2011; Jennings et al, 2015; Ziv et al, 2013). The integrated microscope has a ~0.5 mm 2 field of view (Ghosh et al, 2011), which permits dense sampling of up to ~1,000 individual neurons simultaneously (Alivisatos et al, 2013; Chen et al, 2013a; Ziv et al, 2013), about 20–50 times more cells than can be individually monitored in behaving mice using electrophysiological methods.…”

Visualization of cortical, subcortical and deep brain neural circuit dynamics during naturalistic mammalian behavior with head-mounted microscopes and chronically implanted lenses

“…Moreover, because in vivo electrophysiology relies on waveform shapes to differentiate individual cells from each other, it can be challenging to detect cells with sparse firing patterns or that are located within densely populated networks. Finally, the number of cells that can be detected with in vivo electrophysiology methods is often far less than the number of cells that can be monitored with the optical imaging methods described in this protocol 29,42 . Taken together, these limitations in cell identification and statistical power pose a significant disadvantage for studies that require chronic monitoring of neural activity.…”

Miniature microscopes for large-scale imaging of neuronal activity in freely behaving rodents

“…A recent study provided the first proof-of-principle for this approach. It showed that a systemic administration of Zolpidem, a commonly prescribed in the US GABA-A agonist drug for treating insomnia, reduces hippocampal neuronal activity in freely behaving mice [61]. While such a result may not be surprising, it suggests that comparing neuronal dynamics of drug-treated and control mice in brain disease models may help identify the effects of different molecular factors on network dysfunction and cognitive/behavioral impairments.…”