Shift of intracellular chloride concentration in ganglion and amacrine cells of developing mouse retina.
GABA and glycine provide excitatory action during early development: they depolarize neurons and increase intracellular calcium concentration. As neurons mature, GABA and glycine become inhibitory. This switch from excitation to inhibition is thought to result from a shift of intracellular chloride concentration ([Cl-]i) from high to low, but in retina, measurements of [Cl-]i or chloride equilibrium potential (ECl) during development have not been made. Using the developing mouse retina, we systematically measured [Cl-]i in parallel with GABA's actions on calcium and chloride. In ganglion and amacrine cells, fura-2 imaging showed that before postnatal day (P) 6, exogenous GABA, acting via ionotropic GABA receptors, evoked calcium rise, which persisted in HCO3- -free buffer but was blocked with 0 extracellular calcium. After P6, GABA switched to inhibiting spontaneous calcium transients. Concomitant with this switch we observed the following: 6-methoxy-N-ethylquinolinium iodide (MEQ) chloride imaging showed that GABA caused an efflux of chloride before P6 and an influx afterward; gramicidin-perforated-patch recordings showed that the reversal potential for GABA decreased from -45 mV, near threshold for voltage-activated calcium channel, to -60 mV, near resting potential; MEQ imaging showed that [Cl-]i shifted steeply around P6 from 29 to 14 mM, corresponding to a decline of ECl from -39 to -58 mV. We also show that GABAergic amacrine cells became stratified by P4, potentially allowing GABA's excitatory action to shape circuit connectivity. Our results support the hypothesis that a shift from high [Cl-]i to low causes GABA to switch from excitatory to inhibitory.
Zhang
Ling-Li
LL
Deaprtment of Neuroscience, University of Pennsylvania School of Medicine, Philadelphia, PA 19104-6058, USA.
Pathak
Hemal R
HR
Coulter
Douglas A
DA
Freed
Michael A
MA
Vardi
Noga
N
D000200:Action Potentials / Q000502:physiology; D000375:Aging / Q000502:physiology; D025042:Amacrine Cells / Q000378:metabolism*; D000818:Animals; D000831:Animals, Newborn; D002118:Calcium / Q000378:metabolism / Q000494:pharmacology; D015220:Calcium Channels / Q000502:physiology; D002712:Chlorides / Q000378:metabolism*; D003952:Diagnostic Imaging / Q000379:methods; D016257:Fura-2; D006096:Gramicidin; D008564:Membrane Potentials; D051379:Mice; D008856:Microscopy, Fluorescence / Q000379:methods; D009433:Neural Inhibition / Q000502:physiology; D018408:Patch-Clamp Techniques; D011806:Quinolinium Compounds; D018079:Receptors, GABA / Q000502:physiology; D012160:Retina / Q000254:growth & development* / Q000502:physiology; D012165:Retinal Ganglion Cells / Q000378:metabolism*; D013997:Time Factors; D005680:gamma-Aminobutyric Acid / Q000494:pharmacology / Q000502:physiology
D015220:Calcium Channels; D002712:Chlorides; D011806:Quinolinium Compounds; D018079:Receptors, GABA; C070311:6-methoxy-N-ethylquinolinium; D006096:Gramicidin; D005680:gamma-Aminobutyric Acid; D002118:Calcium; D016257:Fura-2