Chronic in vivo multi-circuit neurophysiological recordings in mice

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Abstract

While genetically modified mice have become a widely accepted tool for modeling the influence of gene function on the manifestation of neurological and psychiatric endophenotypes, only modest headway has been made in characterizing the functional circuit changes that underlie the disruption of complex behavioral processes in various models. This challenge partially arises from the fact that even simple behaviors require the coordination of many neural circuits vastly distributed across multiple brain areas. As such, many independent neurophysiological alterations are likely to yield overlapping circuit disruptions and ultimately lead to the manifestation of similar behavioral deficits. Here we describe the expansion of our neurophysiological recording approach in an effort to quantify neurophysiological activity across many large scale brain circuits simultaneously in freely behaving genetically modified mice. Using this expanded approach we were able to isolate up to 70 single neurons and record local field potential (LFP) activity simultaneously across 11 brain areas. Moreover, we found that these neurophysiological signals remained viable up to 16 months after implantation. Thus, our approach provides a powerful tool that will aid in dissecting the central brain network changes that underlie the complex behavioral deficits displayed by various genetically modified mice.

Research highlights

▶ Whole circuit recordings in mice. ▶ Chronic multi-site implants. ▶ Dynamic oscillatory activity across circuits.

Introduction

Genetically modified (GM) mice have become a widely accepted tool for modeling the influence of gene function on behavior (Ekstrand et al., 2007, Giros et al., 1996, Mohn et al., 1999, Roybal et al., 2007, Welch et al., 2007); however, only modest headway has been made in characterizing the functional changes that underlie the disruption of complex behavioral processes observed across various models. The central challenge lies within the fact that many of the GM mouse lines used to model central nervous system (CNS) disorders display alterations in genes that are expressed across multiple brain areas. Furthermore, even in cases where genetic manipulations are restricted to single brain regions, these targeted manipulations have been shown to induce secondary changes across other brain areas (Kellendonk et al., 2006). Finally, since even simple behaviors require the activation of neural networks which span multiple brain regions, many different isolated brain changes are likely to be sufficient to alter circuit function and ultimately facilitate similar gross behavioral deficits. Taken together, these challenges present a major obstacle in identifying the specific genetic manipulation-induced brain changes that directly underlie the manifestation of a particular set of circuit deficits and ultimately behavioral dysfunction.

In vivo neurophysiological recordings have become an emerging tool for probing the functional molecular components of brain oscillatory networks in various GM mouse models during freely moving behavior (Dzirasa, 2008). For example, genetic disruption of the 5-HT1A receptor potentiates hippocampal theta (4–11 Hz) oscillatory activity during anxiety related task performance (Gordon et al., 2005). Moreover, genetic disruption of the dopamine transporter (DAT) potentiates hippocampal gamma (30–50 Hz) oscillatory activity during exploration of a novel environment (Dzirasa et al., 2006), and diminishes peak hippocampal theta oscillatory frequencies in the home cage and during REM sleep (Dzirasa et al., 2009b). Knock-out (KO) mice for the gap junction protein Connexin-36 have also been shown to display impairments in hippocampal gamma oscillatory coordination (Buhl et al., 2003), and in an elegant gene by drug interaction study, the targeted activation of genetically modified M3 cholinergic receptors has been shown to induce hippocampal gamma oscillatory activity (Alexander et al., 2009). Notably, in vivo recordings have also been utilized to investigate how selective gene manipulations alter neuronal firing rates and firing patterns in freely behaving mice (Adhikari et al., 2010, Alexander et al., 2009, Costa et al., 2006, McHugh et al., 2007, Zweifel et al., 2009).

Since the induction of behavior requires the activation of neural circuits spanning multiple brain regions, an emerging number of studies have been geared towards probing circuit function in GM mice via in vivo neurophysiological recordings conducted simultaneously across multiple brain areas. For example, it has been shown that acute dopamine depletion in mice lacking the dopamine transporter induces the synchronization of cortical–striatal ensembles (Costa et al., 2006). Moreover, studies have shown that mice genetically engineered to display NMDA receptor hypofunction exhibit enhanced hippocampus–prefrontal cortex inter-area theta–gamma coupling, and diminished intra-area theta–gamma coupling (Dzirasa et al., 2009a). More recently, an elegant behavioral study demonstrated that hippocampus and prefrontal cortex theta oscillations synchronize during anxiety related behaviors, and that genetic disruption of the 5-HT1A receptor alters network processing across the hippocampal–prefrontal cortex pathway (Adhikari et al., 2010). Altogether, these studies demonstrate the utility of in vivo recordings in probing circuit deficits underling the behavioral manifestations seen in various lines of transgenic mice, and highlight the need for the development of tools which allow investigators to record more neurons per mouse and neural activity across entire brain circuits concurrently.

Section snippets

Animal care and use

Mice were separated into individual cages maintained in a humidity and temperature-controlled room with water available ad libitum, and surgically implanted with recording electrodes at 2–5 months. Recordings were initiated following a 2–4 week recovery period. All neurophysiological recordings were conducted in an 11″ × 11″ open field test environment.

All studies were conducted with approved protocols from the Duke University Institutional Animal Care and Use Committee and were in accordance

Results

In order to increase the number of neurons that we could acquire from each mouse, we set out double the number implanted microwires to 64. Our initial approach entailed implanting four multiwire electrode arrays (16-microwire arrays) in each mouse. While we have used this approach successfully in the past to increase the number of neurons isolated in rats and non-human primates (Nicolelis et al., 1997, Nicolelis et al., 2003), we found that this approach did not work in mice. Each implanted

Discussion

GM mice have become a widely accepted tool for modeling the influence of gene function on behavior (Ekstrand et al., 2007, Giros et al., 1996, Mohn et al., 1999, Roybal et al., 2007, Welch et al., 2007); however, only modest headway has been made in characterizing the functional brain changes that underlie the disruption of complex behavioral processes observed across various models. Since these behavioral deficits ultimately result from changes in neural circuits which span multiple brain

Conclusion

Overall, our chronic neurophysiological recording approach provides a powerful tool for dissecting the circuit changes underlying behavioral changes seen various GM mouse models of neurological and psychiatric diseases and probing the effect of pharmacological agents on these circuits. Moreover, our recording approach can potentially be integrated with novel optogenetic and chemicogenetic tools capable of manipulating neuronal activity in order to quantify the role that specific cell types play

Acknowledgements

We would like to thank L. Oliveira, T. Jones, and G. Wood for miscellaneous support, and S. Halkiotis for proofreading this manuscript. This work was supported by funding from UNCF/Merck, and NIMH grant P50MH060451-09S1 to KD; and by NIH grant R33NSO49534, and The Safra Foundation to MALN. A special thanks to Freeman Hrabowski, Robert and Jane Meyerhoff, and the Meyerhoff Scholarship Program.

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