Calcium dependence of differentiation of GABA immunoreactivity in spinal neurons

Abstract

The developmental regulation of neurotransmitter synthesis has been extensively studied and appears in many cases to depend on electrical activity. The central nervous system of the Xenopus embryo and young larva is an attractive subject for such studies, since action potentials first elicited from Xenopus spinal neurons at the time of closure of the neural tube are long in duration and calcium-dependent. Moreover, cells exhibit spontaneous elevations of intracellular calcium during this early period as a consequence of calcium influx through voltage-dependent channels, which induces calcium release from intracellular stores. Since the early differentiation of Xenopus spinal neurons in dissociated cell culture parallels development in vivo, we have examined the maturation of gamma-aminobutyric acid (GABA) immunoreactivity in cultured neurons and explored its dependence on spontaneous calcium influx at early stages of development. We find that specific GABA immunoreactivity develops in spinal neurons in dissociated cell culture with the same time course previously defined in vivo. Additionally, this process requires calcium influx that occurs spontaneously through voltage-dependent channels. The appearance of GABA immunoreactivity is blocked by transcriptional inhibitors. The early appearance of GABA raises the possibility that it may play additional roles at early stages of development.

GABA: A Pioneer Transmitter That Excites Immature Neurons and Generates Primitive Oscillations

“…These early effects are briefly summarized here as they provide an excellent illustration of the multiple facets of the actions of GABA and the unique role of GABA as an early communicating signal. Excellent reviews of these effects have been published (36,352,492,579).…”

Ion Channel Development, Spontaneous Activity, and Activity-Dependent Development in Nerve and Muscle Cells

“…When spikes and waves are eliminated in 0 Ca 2ϩ medium, the normal speeding of delayed K ϩ current activation does not occur, fewer neurons develop the GABA phenotype (quantified by both GABA immunoreactivity and GAD transcripts), and neurites are abnormally long (145, 248,249,557,607).…”

Onset of Electrical Excitability during a Period of Circus Plasma Membrane Movements in DifferentiatingXenopusNeurons

“…Xenopus spinal neurons cultured from neurula stage embryos exhibit spontaneous, transient elevations of intracellular calcium ([Ca 2ϩ ] i ) before and after neurite initiation that are required for appropriate expression of the neurotransmitter GABA and maturation of K ϩ current kinetics (Holliday and Spitzer, 1990;Desarmenien and Spitzer, 1991;Spitzer et al, 1993;Gu and Spitzer, 1995). By the earliest time of neurite initiation, primary spinal neurons display Ca 2ϩ -dependent action potentials and mature Ca 2ϩ current density (Spitzer and Lamborghini, 1976;Barish, 1986;O'Dowd et al, 1988;Spitzer, 1991), implying that voltagedependent currents appear at an earlier stage.…”

“…Previous work demonstrated that Ca 2ϩ influx through voltagedependent Ca 2ϩ channels regulates aspects of spinal neuron differentiation (Desarmenien and Spitzer, 1991;Spitzer et al, 1993;Gu and Spitzer, 1995). Because voltage-clamped Ca 2ϩ currents in circus cells were typically small, Ca 2ϩ imaging was used to determine whether they are sufficient to produce detectable elevations of [Ca 2ϩ ] i in response to depolarization and to confirm the developmental profile of expression of Ca 2ϩ currents during the first 6 hr in culture.…”