Depolarization promotes GAD 65‐mediated GABA synthesis by a post‐translational mechanism in neural stem cell‐derived neurons

Abstract

Neuronal activity regulates neurogenesis and neuronal differentiation in the mammalian brain. The commencement of neurotransmitter expression establishes the neuronal phenotype and enables the formation of functional connectivity between neurons. In addition, release of neurotransmitters from differentiating neurons may modulate the behaviour of neural precursors. Here, we show that neuronal activity regulates gamma-aminobutyric acid (GABA) expression in neurons generated from stem cells of the striatum and adult subventricular zone (SVZ). Differentiating neurons display spontaneous Ca2+ events, which are voltage-gated calcium channel (VGCC) dependent. Depolarization increases both the frequency of Ca2+ transients and the amount of Ca2+ influx in differentiating neurons. We show that depolarization-dependent GABA expression is regulated by the amplitude and not by the frequency of Ca2+ influx. Brief activation of VGCCs leads to Ca2+ influx that in turn promotes a rapid expression of GABA. Depolarization-dependent GABA expression does not require changes in gene expression. Instead, it involves cAMP-dependent protein kinase (PKA) and Ca2+ and phospholipid-dependent protein kinase (PKC) signalling. Activity increases the number of glutamic acid decarboxylase (GAD) 65-immunoreactive neurons in a PKA-dependent manner, without altering the expression of GAD 65, suggesting that depolarization promotes recruitment of GAD 65 by a post-translational mechanism. In line with this, depolarization does not permanently increase the expression of GABA in neurons derived from neural stem cells of the embryonic striatum, cortex and adult SVZ. Thus, neuronal activity does not merely accelerate neuronal differentiation but it may alter the mechanism of GABA synthesis in newly generated neurons.

Adult neurogenesis and brain regeneration in zebrafish

“…Several other regulators of stem cells in the adult vertebrate brain have been implicated, such as Nitric oxide (Gibbs, 2003;Moreno-López and González-Forero, 2006), glutamate and c-aminobutryic acid (Schlett, 2006;Jagasia et al, 2006;Gakhar-Koppole, 2008;Markwardt and Overstreet-Wadiche, 2008;Mattson, 2008;Platel et al, 2010), dopamine (Cave and Baker, 2009), vascular endothelial growth factor (VEGF) (Jin et al, 2002;Sun et al, 2003;Zhu et al, 2003;Sun and Guo, 2005;Wittko et al, 2009), pigment epithelium derived growth factor (PEDF) (Ramirez-Castillejo et al, 2006), glial-cell derived neurotrophic factor (GDNF) (Chen et al, 2005;Kobayashi et al, 2006;Wei et al, 2007), brain-derived neurotrophic factor (BDNF) (Benraiss et al, 2001;Pencea et al, 2001;Scharfman et al, 2005;Rossi et al, 2006;Chan et al, 2008;Lee and Son, 2009), and epigenetic modifications by histone methylation , adding further degrees of complexity to the regulation of adult stem cells in the vertebrate brain. The roles of these regulators in zebrafish brain have not been addressed, which offers an interesting research realm.…”

GABA Not Only a Neurotransmitter: Osmotic Regulation by GABAAR Signaling

“…Consistently, GABA evoked a rapid Ca 2+ increase in neuroblasts but not in precursor cells (Cesetti et al, 2010). However, spontaneous Ca 2+ signals occur in vitro both in embryonic (Ciccolini et al, 2003; Gakhar-Koppole et al, 2008) and in neonatal neural progenitors (Cesetti et al, 2009) and they can be modulated by GABA (Figure 3). GABAergic modulation of spontaneous Ca 2+ signals was also observed in situ in GFAP expressing cells of the SVZ; these spontaneous Ca 2+ signals depended on IP 3 mediated Ca 2+ release from the stores.…”

GABAA Receptor Signaling Induces Osmotic Swelling and Cell Cycle Activation of Neonatal Prominin+ Precursors

“…The SEZ of the lateral ventricle was dissected and dissociated cells were processed for sorting either immediately or after being plated overnight at a density of 10 5 cells per ml in NS-A (Euroclone, Eching, Germany) medium supplemented with 10 ng/ml fibroblast growth factor (FGF)-2 (R&D System, Wiesbaden-Nordenstadt, Germany) as already described [10]. Proliferation medium contained NS-A or Neurobasal (NBA; Invitrogen, Darmstadt, Germany) media, 2% B27, 2 mM Lglutamine, penicillin 100 U/ml, streptomycin 100 mg/ml, FGF-2 10 ng/ml, and EGF 20 ng/ml.…”

“…For Ca 2þ imaging, the cells were loaded at room temperature (RT) for 20 minutes with 2-4 lM Fluo3-AM (Molecular Probes) and imaged in Ringer's solution as previously described [10]. For imaging with a potentiometric dye, the cells were stained at RT for 10 minutes with 75 nM DiBAC 4 [3] (Molecular Probes) in Ringer's solution and imaged in the same solution.…”