Interactions between NMDA receptors and mGlu5 receptors expressed in HEK293 cells

1 Ca2þ imaging was used to investigate interactions between responses induced by N-methyl-D-aspartate (NMDA; 15 mM) and (RS)-3,5-dihydroxyphenyl-glycine (DHPG; 30mM) in human embryonic kidney (HEK) 293 cells, transiently transfected with rat recombinant NR1a, NR2A and mGlu5a cDNA. 2 Responses to NMDA were reversibly depressed by DHPG from 244714 to 194712 % of baseline. Treatment with thapsigargin (1 mM, 10min) prevented this effect. 3 After thapsigargin pretreatment, repeated applications of NMDA showed a gradual rundown in amplitude over a period of several hours, and were unaffected by DHPG. 4 Continuous perfusion with staurosporine (0.1 mM), after thapsigargin pretreatment, converted the run-down to a small increase in NMDA responses to 12376 % of baseline. DHPG induced a further and sustained potentiation of NMDA responses to 174712 % of the initial baseline. 5 The protein tyrosine kinase (PTK) inhibitors genistein (50 mM) and 3-(4-chlorophenyl)1-(1,1-dimethylethyl)-1H-pyrazolo[3,4-d]pyrimidin-4-amine (PP2; 1 mM) inhibited the staurosporine- and DHPG-induced potentiation of NMDA responses. 6 The protein phosphatase (PTP) inhibitors orthovanadate (100 mM) and phenyl arsine oxide (PAO