Rac1 and Rac3 GTPases and TPC2 are required for axonal outgrowth and migration of cortical interneurons.

Rho GTPases, among them Rac1 and Rac3, are major transducers of extracellular signals and are involved in multiple cellular processes. In cortical interneurons, the neurons that control the balance between excitation and inhibition of cortical circuits, Rac1 and Rac3 are essential for their development. Ablation of both leads to a severe reduction in the numbers of mature interneurons found in the murine cortex, which is partially due to abnormal cell cycle progression of interneuron precursors and defective formation of growth cones in young neurons. Here, we present new evidence that upon Rac1 and Rac3 ablation, centrosome, Golgi complex and lysosome positioning is significantly perturbed, thus affecting both interneuron migration and axon growth. Moreover, for the first time, we provide evidence of altered expression and localization of the two-pore channel 2 (TPC2) voltage-gated ion channel that mediates Ca2+ release. Pharmacological inhibition of TPC2 negatively affected axonal growth and migration of interneurons. Our data, taken together, suggest that TPC2 contributes to the severe phenotype in axon growth initiation, extension and interneuron migration in the absence of Rac1 and Rac3.

Cortical interneuron development: a role for small Rho GTPases.

GABAergic interneurons control cortical excitation/inhibition balance and are implicated in severe neurodevelopmental disorders. In contrast to the multiplicity of signals underlying the generation and migration of cortical interneurons, the intracellular proteins mediating the response to these cues are largely unknown. We have demonstrated the unique and diverse roles of the Rho GTPases Rac1 and 3 in cell cycle and morphology in transgenic animals where Rac1 and Rac1/3 were ablated specifically in cortical interneurons. In the Rac1 mutant, progenitors delay their cell cycle exit, probably due to a prolonged G1 phase resulting in a cortex with 50% reductions in interneurons and an imbalance of excitation/inhibition in cortical circuits. This disruption in GABAergic inhibition alters glutamatergic function in the adult cortex, which could be reversed by enhancement of GABAergic functions during an early postnatal period. Furthermore, this disruption disturbs neuronal synchronization in the adult cortex. In the double mutant, additional severe cytoskeletal defects result in an 80% interneuron decrease. Both lines die postnatally from epileptic seizures. We have made progress towards characterizing the cell cycle defect in Rac1 mutant interneuron progenitors, determining the morphological and synaptic characteristics of single and double mutant interneurons and identifying some of the molecular players through which Racs exert their actions via proteomic analysis. In our present work, we review these studies and discuss open questions and future perspectives. We hope that our data will contribute to the understanding of the function of cortical interneurons, especially since preclinical models of interneuron-based cell therapies are being established.

Rac-GTPases Regulate Microtubule Stability and Axon Growth of Cortical GABAergic Interneurons.

Cortical interneurons are characterized by extraordinary functional and morphological diversity. Although tremendous progress has been made in uncovering molecular and cellular mechanisms implicated in interneuron generation and function, several questions still remain open. Rho-GTPases have been implicated as intracellular mediators of numerous developmental processes such as cytoskeleton organization, vesicle trafficking, transcription, cell cycle progression, and apoptosis. Specifically in cortical interneurons, we have recently shown a cell-autonomous and stage-specific requirement for Rac1 activity within proliferating interneuron precursors. Conditional ablation of Rac1 in the medial ganglionic eminence leads to a 50% reduction of GABAergic interneurons in the postnatal cortex. Here we examine the additional role of Rac3 by analyzing Rac1/Rac3 double-mutant mice. We show that in the absence of both Rac proteins, the embryonic migration of medial ganglionic eminence-derived interneurons is further impaired. Postnatally, double-mutant mice display a dramatic loss of cortical interneurons. In addition, Rac1/Rac3-deficient interneurons show gross cytoskeletal defects in vitro, with the length of their leading processes significantly reduced and a clear multipolar morphology. We propose that in the absence of Rac1/Rac3, cortical interneurons fail to migrate tangentially towards the pallium due to defects in actin and microtubule cytoskeletal dynamics.