Disrupted GABAAR trafficking and synaptic inhibition in a mouse model of Huntington's disease

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Abstract

Growing evidence suggests that Huntington's disease (HD), a neurodegenerative movement disorder caused by the mutant huntingtin (htt) with an expanded polyglutamine (polyQ) repeat, is associated with the altered intracellular trafficking and synaptic function. GABAA receptors, the key determinant of the strength of synaptic inhibition, have been found to bind to the huntingtin associated protein 1 (HAP1). HAP1 serves as an adaptor linking GABAA receptors to the kinesin family motor protein 5 (KIF5), controlling the transport of GABAA receptors along microtubules in dendrites. In this study, we found that GABAAR-mediated synaptic transmission is significantly impaired in a transgenic mouse model of HD expressing polyQ-htt, which is accompanied by the diminished surface expression of GABAA receptors. Moreover, the GABAAR/HAP1/KIF5 complex is disrupted and dissociated from microtubules in the HD mouse model. These results suggest that GABAAR trafficking and function is impaired in HD, presumably due to the interference of KIF5-mediated microtubule-based transport of GABAA receptors. The diminished inhibitory synaptic efficacy could contribute to the loss of the excitatory/inhibitory balance, leading to increased neuronal excitotoxicity in HD.

Highlights

► We compared synaptic inhibition in wild-type and a transgenic mouse model of HD. ► We examined GABAA receptor surface expression in the HD mouse model. ► We examined the GABAAR/KIF5/microtubule complex in the HD mouse model. ► We find that GABAAR-mediated synaptic inhibition is impaired in HD. ► We find that KIF5-mediated microtubule-based transport of GABAARs is disturbed in HD.

Introduction

Huntington's disease (HD) is a devastating neurological disorder characterized by uncontrolled movements, which is associated with the dysfunction and eventually degeneration of striatal medium spiny neurons (MSNs, Vonsattel and DiFiglia, 1998, Vonsattel et al., 1985). The GABAergic MSNs account for > 90% neuronal population in the striatum, a key area in basal ganglia whose main function is the movement control. Genetic studies have found that HD is caused by an abnormally elongated polyglutamine (polyQ) tract in the large protein huntingtin (htt, Mangiarini et al., 1996), however, both the normal function of htt in neurons and the molecular mechanism by which the expanded polyQ sequence in htt causes selective neurodegeneration remain elusive. In addition to roles in regulating apoptosis and transcription (Ross, 2002), polyQ-htt may in part mediate its neurotoxic action in HD by altering neuronal membrane trafficking and synaptic function (Fan and Raymond, 2007, Smith et al., 2005). Several htt-interacting proteins implicated in intracellular transport have been identified (Harjes and Wanker, 2003), one of which is huntingtin-associated protein 1 (HAP1) (Li et al., 1995). HAP1 interacts more tightly with polyQ-htt than wild-type htt (Li et al., 1995), and may act as a key mediator of pathological alterations in membrane trafficking by mutant htt (Gauthier et al., 2004, Li and Li, 2005, Rong et al., 2006).

HAP1 associates with kinesin or dynein microtubule motor proteins (Engelender et al., 1997, Gauthier et al., 2004, Li et al., 1998, McGuire et al., 2006, Twelvetrees et al., 2010). Growing evidence suggests that mutant huntingtin impairs the HAP1/motor-dependent anterograde or retrograde transport of neuronal cargos along microtubules (Gauthier et al., 2004, Rong et al., 2006). Deficits in these neuronal transport systems have been suggested to underlie the pathogenesis of a number of neurodegenerative diseases (Goldstein, 2003).

Fast inhibitory neurotransmission mediated by GABAA receptors (GABAARs) plays a critical role in regulating neuronal excitability. The trafficking of GABAARs underlies dynamic changes in synaptic receptor numbers and inhibitory postsynaptic current amplitudes, providing an effective mechanism for regulating the strength and plasticity of synaptic inhibition (Jacob et al., 2008). One critical determinant for GABAAR trafficking and inhibitory transmission is the kinesin family member KIF5 motor protein, which associates with HAP1 (Twelvetrees et al., 2010). Based on studies in transfected neuronal cultures, it has been found that HAP1 interacts with GABAARs, facilitating the recycling of internalized GABAARs back to synapses (Kittler et al., 2004), and suppressing HAP1 expression attenuates GABAAR trafficking and synaptic inhibition (Twelvetrees et al., 2010). In this study, we sought to determine whether synaptic inhibition is impaired in a mouse model of HD, and whether it results from the loss of GABAAR transport along microtubules due to the disruption of the HAP1/KIF5/GABAAR multiprotein complex in vivo. The diminished strength of inhibitory synaptic transmission could contribute to the loss of the excitatory/inhibitory balance, leading to increased neuronal excitotoxicity.

Section snippets

Animals

All experiments were performed with the approval of State University of New York at Buffalo Animal Care Committee. The transgenic mouse model of HD, N171-82Q, which expresses a mutant N-terminal fragment of huntingtin (the first 171 aa of human htt with 82Q, Schilling et al., 1999), was purchased from Jackson Lab. Experiments were conducted at the symptomatic stage (3–5 months old) unless otherwise stated.

Electrophysiological recordings in slices

Mice were first anesthetized by inhaling Halothane (Sigma) for ~ 30 s and decapitated

GABAAR-mediated inhibitory transmission is disrupted in HD

Since GABAAR-mediated synaptic response is impaired by transfected polyQ-htt in neuronal cultures (Twelvetrees et al., 2010), we hypothesize that mice with in vivo expression of mutant huntingtin might show altered GABAergic transmission. To test this, we examined GABAAR-mediated inhibitory postsynaptic current (IPSC) in a transgenic mouse model of HD, N171-82Q, which expresses a mutant N-terminal fragment of huntingtin. N171-82Q mice develop behavioral abnormalities resembling HD, including

Discussion

It is known that GABAergic transmission plays a key role in the neuronal communication in striatal or cortical circuits. GABAAR dysfunction is implicated in multiple neurological diseases, such as epilepsy, anxiety disorders, fragile X syndrome, and schizophrenia (Benarroch, 2007, D'Hulst and Kooy, 2007, Lewis and Gonzalez-Burgos, 2006, Rudolph and Möhler, 2004). In HD studies, GABAergic transmission in cortical and striatal neurons has been examined in different mouse models, such as R6/2 mice

Conclusion

In summary, our results show that the HAP1/KIF5-mediated anterograde transport of GABAARs along dendritic microtubules is impaired by the mutant htt in HD conditions. PolyQ-htt alters GABAAR vesicle transport, resulting in reduced surface delivery and accumulation of GABAARs at inhibitory synapses and ensuing reduced inhibitory synaptic response. Blocking polyQ-htt disruption of the machinery underlying HAP1/KIF5-facilitated trafficking of GABAARs to synapses may be a therapeutic approach for

Acknowledgments

We thank Xiaoqing Chen for excellent technical support. This work was supported by NIH R21 grant NS069929 to Z.Y.

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    These authors contributed equally to this work.

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