Cross talk between the GABA(A) receptor and the Na-K-Cl cotransporter is mediated by intracellular Cl-.
It has been suggested that the GABA(A) receptor-mediated depolarization in immature neurons depends on a high intracellular Cl(-) concentration maintained by Na-K-Cl cotransporter isoform1 (NKCC1). We previously found that activation of the GABA(A) receptor led to stimulation of NKCC1. This stimulation could be a result of GABA(A) receptor-mediated Cl(-) efflux. However, a loss of intracellular Cl(-) is associated with cell shrinkage, membrane depolarization, as well as a rise of intracellular Ca(2+) concentration ([Ca(2+)](i)). To determine which cellular mechanism is underlying NKCC1 stimulation, we investigated changes of intracellular Cl(-) content, [Ca(2+)](i), cell volume, and NKCC1 activity following GABA(A) receptor activation. The basal levels of intracellular (36)Cl were 0.70 +/- 0.04 micromol/mg protein. The intracellular (36)Cl content decreased to 0.53 +/- 0.03 micromol/mg protein in response to 30 microM muscimol (P < 0.05). The loss of intracellular (36)Cl was blocked by 10 microM bicuculline. Muscimol triggered a rise in [Ca(2+)](i), but did not cause cell shrinkage. In contrast, 10-50 mM [Cl(-)](o) or hypertonic HEPES-MEM resulted in reversible cell shrinkage (P < 0.05). Moreover, the GABA-mediated stimulation of NKCC1 activity was not abolished either by removal of extracellular Ca(2+) or BAPTA-AM. An increase in phosphorylation of NKCC1 was detected under both 10 mM [Cl(-)](o) and muscimol conditions. These results suggest that a GABA-mediated loss of intracellular Cl(-), but not a subsequent rise in [Ca(2+)](i) or shrinkage, leads to stimulation of NKCC1. This stimulation may be an important positive feedback mechanism to maintain intracellular Cl(-) level and GABA function in immature neurons.
Schomberg
Stacey L
SL
Department of Neurological Surgery, University of Wisconsin Medical School, Madison 53792, USA.
Bauer
James
J
Kintner
Douglas B
DB
Su
Gui
G
Flemmer
Andreas
A
Forbush
Biff
B
Sun
Dandan
D
D000818:Animals; D002118:Calcium / Q000378:metabolism; D048429:Cell Size; D002478:Cells, Cultured; D002540:Cerebral Cortex / Q000166:cytology / Q000196:embryology; D002712:Chlorides / Q000378:metabolism*; D005260:Female; D018755:GABA Agonists / Q000494:pharmacology; D009118:Muscimol / Q000494:pharmacology; D009474:Neurons / Q000166:cytology / Q000378:metabolism*; D010766:Phosphorylation; D011188:Potassium / Q000378:metabolism; D011247:Pregnancy; D051381:Rats; D017207:Rats, Sprague-Dawley; D020239:Receptor Cross-Talk / Q000502:physiology*; D011963:Receptors, GABA-A / Q000378:metabolism*; D028021:Sodium-Potassium-Chloride Symporters / Q000378:metabolism*; D064506:Solute Carrier Family 12, Member 2
D002712:Chlorides; D018755:GABA Agonists; D011963:Receptors, GABA-A; C578879:Slc12a2 protein, rat; D028021:Sodium-Potassium-Chloride Symporters; D064506:Solute Carrier Family 12, Member 2; D009118:Muscimol; D011188:Potassium; D002118:Calcium