Academic journal article Genetics

Pharmacogenetic Analysis Reveals a Post-Developmental Role for Rac GTPases in Caenorhabditis Elegans GABAergic Neurotransmission

Academic journal article Genetics

Pharmacogenetic Analysis Reveals a Post-Developmental Role for Rac GTPases in Caenorhabditis Elegans GABAergic Neurotransmission

Article excerpt

ABSTRACT

The nerve-cell cytoskeleton is essential for the regulation of intrinsic neuronal activity. For example, neuronal migration defects are associated with microtubule regulators, such as LIS1 and dynein, as well as with actin regulators, including Rac GTPases and integrins, and have been thought to underlie epileptic seizures in patients with cortical malformations. However, it is plausible that post-developmental functions of specific cytoskeletal regulators contribute to the more transient nature of aberrant neuronal activity and could be masked by developmental anomalies. Accordingly, our previous results have illuminated functional roles, distinct from developmental contributions, for Caenorhabditis elegans orthologs of LIS1 and dynein in GABAergic synaptic vesicle transport. Here, we report that C. elegans with function-altering mutations in canonical Rac GTPase-signaling-pathway members demonstrated a robust behavioral response to a GABA^sub A^ receptor antagonist, pentylenetetrazole. Rac mutants also exhibited hypersensitivity to an acetylcholinesterase inhibitor, aldicarb, uncovering deficiencies in inhibitory neurotransmission. RNA interference targeting Rac hypomorphs revealed synergistic interactions between the dynein motor complex and some, but not all, members of Rac-signaling pathways. These genetic interactions are consistent with putative Rac-dependent regulation of actin and microtubule networks and suggest that some cytoskeletal regulators cooperate to uniquely govern neuronal synchrony through dynein-mediated GABAergic vesicle transport in C. elegans.

EPILEPSY affects 1-2% of the worldpopulationandis associated with imbalances between excitatory and inhibitory neurotransmission in the brain (Locke et al. 2009). In particular, interneurons expressing gammaaminobutyric acid (GABA), the principal inhibitory neurotransmitter in the human brain, are essential for normal neuronal synchronization and maintenance of a seizure threshold in humans (Cossette et al. 2002), rodents (Delorey et al. 1998), and zebrafish (Baraban et al. 2005). A failure of the brain to properly regulate neuronal synchrony can result from ion channel defects (Xu and Clancy 2008), neuropeptide depletion (Brill et al. 2006), brain malformations (Patel et al. 2004), interneuron loss (Cobos et al. 2005), and/or synaptic vesicle recycling failure (Di Paolo et al. 2002), all of which may be caused by disrupting the nerve-cell cytoskeleton. Therefore, further exploration of putative links between cytoskeletal components and neurotransmission may accelerate development of novel therapeutics for epilepsy.

Epilepsy associated with cytoskeletal dysfunction often has a developmental basis (Di Cunto et al. 2000; Wenzel et al. 2001; Keays et al. 2007). For example, mutations in LIS1, a dynein motor complex regulator, lead to classical lissencephaly, which is characterized by neuronal migration defects, a lack of convolutions in the brain, mental retardation, and epileptic seizures (Lo Nigro et al. 1997). Yet, observations that lissencephalyassociated seizures worsen after neuronal migration ceases, while LIS1 expression persists, imply that LIS1 also acts in the adult brain (Cardoso et al. 2002).

We previously reported that C. elegans with a predicted null mutation (t1550) in lis-1, the worm ortholog of human LIS1, exhibited synaptic vesicle misaccumulations, but not neuronal migration or axon-pathfinding defects, in GABAergic motor neurons.We also observed anterior "epileptic-like" convulsions, which were intense, frequent, and repetitive, with lis-1(t1550) homozygotes in the presence of pentylenetetrazole (PTZ; Williams et al. 2004), an epileptogenic GABAA receptor antagonist (Huang et al. 2001; Fernandez et al. 2007). PTZ sensitivity was also increased in heterozygous lis-1(t1550) mutants following RNA interference (RNAi) against worm orthologs of associated cortical malformation genes, such as cdk-5 and nud-2, which are known to interact with LIS1 and the dynein motor complex. …

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