[Ca-dependent modulation of human glycine receptors expressed in cultured cell lines].
Glycine receptors (GlyRs) provide the main inhibitory neurotransmission in spinal cord and brainstem synapses of vertebrates. Fucile et al. (2000) discovered that elevation of intracellular Ca2+ caused rapid potentiation of GlyRs. This modulation develops in less than 100 ms. It is characterized by an increase in GlyR apparent affinity for glycine. It has been suggested that the phenomenon of Ca-induced potentiation involves an unknown Ca2+-binding protein (CaBP). Using the yeast two-hybrid system, screening of human brain cDNA library against the cytoplasmic loop of human alpha 1 subunit (GlyRhl) allowed us to identify five new interactors. One of them belongs to a family of Ca-binding proteins. We analyzed effect of "short" forms of this protein (CaBP-S) on functional properties of GlyRhl expressed in HEK-293 and CHO cells. Using whole-cell recordings and rapid agonist application we constructed concentration dependencies of glycine-induced currents. This analysis revealed statistical differences in EC50s between control cells (expressing only GlyRhl) and those expressing CaBP-S. In HEK-293 cells recorded under conditions of low intracellular Ca concentration (BAPTA 20 mM in the recording pipette), EC50 for glycine in control cells and expressing GlyRhl + CaBP-S were, correspondently, 68+/-49 microM (n = 29) and 409 +/-421 microM (n = 60). In CHO cells EC50 were 54+/-43 microM (n = 25) and 123 +/-104 microM (n = 28). These differences were statistically not significant at recording with intracellular solution containing high Ca concentration (50 microM). In this case EC50 were correspondently 35+/-28 microM (n = 7) and 64 +/-38 microM (n = 7). These results suggest that CaBP-S causes decrease of GlyR sensitivity to agonist through interaction with cytoplasmic domain of GlyR.
Buldakova
S
S
Real
E
E
Jakob
Y
Y
Brezhestovskiĭ
P
P
D000818:Animals; D016466:CHO Cells; D015220:Calcium Channels; D002135:Calcium-Binding Proteins / Q000378:metabolism*; D045744:Cell Line, Tumor; D006224:Cricetinae; D003412:Cricetulus; D003593:Cytoplasm / Q000378:metabolism; D004553:Electric Conductivity; D006801:Humans; D011485:Protein Binding; D018009:Receptors, Glycine / Q000378:metabolism*
D015220:Calcium Channels; D002135:Calcium-Binding Proteins; D018009:Receptors, Glycine