Role of Bicarbonate and Chloride in GABA- and Glycine-Induced Depolarization and [Ca²⁺]_iRise in Fetal Rat MotoneuronsIn Situ

Abstract

Ca2ϩ imaging and (perforated) patch recording were used to analyze the mechanism of GABA-and glycine-induced depolarizations in lumbar motoneurons of spinal cord slices from fetal rats. In fura-2 ester-loaded cells, the agonist-induced depolarizations increased [Ca 2ϩ ] i by up to 100 nM. The GABA-and glycine-evoked [Ca 2ϩ ] i transients were suppressed by bicuculline and strychnine, respectively. Their magnitude decreased by ϳ50% between embryonic days 15.5 and 19.5. Ϫ efflux has a major contribution to depolarizations mediated by GABA A and glycine receptor-coupled anion channels in prenatal neurons. We hypothesize that the HCO 3 Ϫ -dependent depolarizing component, which is likely to produce an intracellular acidosis, might play an important role during the early postnatal period when the Cl Ϫ -dependent component gradually shifts to hyperpolarization. Key words: bicarbonate; calcium; chloride pump; development; imaging; motor neurons; neuronal maturationThe principal hyperpolarizing and thus inhibitory neurotransmitters GABA and glycine exert a depolarizing action during development of neuronal structures (Ben-Ari et al., 1989;Obrietan and van den Pol, 1995). In the immature hippocampus, depolarizing GABAergic IPSPs inhibit synaptic responses of CA3 pyramidal neurons (Psarropoulou and Descombes, 1999;Palva et al., 2000), but periodic GABA release produces a "giant" neuronal depolarization and action potential discharge (Ben-Ari et al., 1989). The rise in the concentration of free intracellular Ca 2ϩ ([Ca 2ϩ ] i ) associated with the GABA-induced depolarization (Leinekugel et al., 1995;Garaschuk et al., 1998) might be implicated in trophic or hebbian modulation of developing synapses and activity-dependent formation of the hippocampal network Leinekugel et al., 1999).In motoneurons that are among the earliest neurons to differentiate within the brain, activity-related rises of [Ca 2ϩ ] i are also supposed to have a trophic effect. Suppression of neurite outgrowth in developing motoneurons (Owen and Bird, 1997; Metzger et al., 1998) appears to be causally related with an increase of [Ca 2ϩ ] i attributable to activation of Ca 2ϩ -permeable glutamate receptors (Metzger et al., 2000). Accordingly, maturation of motoneurons of cultured lumbar spinal cord is retarded upon blockade of glutamatergic neurotransmission (Xie and Ziskind-Conhaim, 1995). One week before birth, the isolated spinal cord of rats generates rhythmic nerve activity that is impaired by blockers of glycine and GABA A receptors (Nishimaru et al., 1996). It was found previously that GABA and glycine depolarize lumbar motoneurons that provide the output of this rhythmically active network in the fetus (Obata et al., 1978;Wu et al., 1992;Gao and Ziskind-Conhaim, 1995). The latter studies indicated a major role of Cl Ϫ ions in this depolarization. However, it is yet not clear whether efflux of HCO 3 Ϫ through the receptor-coupled anion pore (Bormann et al., 1987;Fatima-Shad and Barry, 1993) contributes to the GABA-and glycine-evoked r...

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

“…This effect is mainly due to the activation of voltage-gated calcium ] i response reversed rapidly upon removal of the transmitter. In keeping with these results, GABA and glycine increased [Ca 2ϩ ] i in a number of embryonic and neonatal neurons including neocortex (223,385,399,675), hippocampus (222,377,378), spinal cord (359), dorsal horn neurons (515,653), hypothalamus, olfactory bulb, cortex, medulla, striatum, thalamus, hippocampus, and colliculus (484) (see Table 1). …”

Increasing the Frequency of Spontaneous Rhythmic Activity Disrupts Pool-Specific Axon Fasciculation and Pathfinding of Embryonic Spinal Motoneurons

“…1 A), as assessed in the in vitro spinal-cord hindlimb preparations prepared from the treated embryos. The increase in episode frequency presumably resulted from preventing the reuptake of endogenously released glycine, which is excitatory at early stages of chick and mouse development (Wu et al, 1992;Nishimaru et al, 1996;Milner and Landmesser, 1999;Kulik et al, 2000;Hanson and Landmesser, 2003) and during the pathfinding events described here, which occur between St 22 and St 28.…”

Use-Dependent Shift From Inhibitory to Excitatory GABAA Receptor Action in SP-O Interneurons in the Rat Hippocampal CA3 Area

“…The second factor was shifts in the transmembrane ion gradients responsible for E GABA-A ; this was shown here experimentally by using formate to alter the intracellular anion load (see Lamsa and Kaila 1997). Weak-acid anions have previously been used in brain slices to study changes in E GABA-A (Kaila et al 1993;Kulik et al 2000;Lamsa and Kaila 1997;see Grover et al 1993;Perkins 1999;Staley et al 1995). As shown by the experiments with formate, the set point and dynamics of the GABA A reversal potential are critical factors in determining the behavior of the CA3 interneuronal network.…”