Figure 4.
Effect of various pharmacological agents on GPCR-mediated [Ca2+]i increase. A, Effect of mGluR agonist or antagonist on AP-induced [Ca2+]i increase at the soma of hippocampal pyramidal neuron filled with the low-affinity indicator fura-6F (Kd, 6 μm) in wild-type mice. Time-lapse imaging of fluorescence changes (top) and voltage responses induced by APs at 30 Hz (bottom) were taken before (−DHPG) and during (+DHPG) bath application of 30 μm DHPG, a group I mGluR agonist. DHPG-induced [Ca2+]i increase in the presence of 1 mm MCPG, a mGluR antagonist (MCPG+DHPG), after washing out of MCPG (+DHPG), and without the presence of any agents (wash). B, Effect of a cholinergic agonist, CCh, on AP-induced [Ca2+]i increase. Time-lapse imaging of fluorescence changes (top) and voltage responses induced by APs at 30 Hz (bottom) were taken before (−CCh) and during (+CCh) bath application of 30 μm CCh. After application of a muscarinic receptor antagonist, atropine (1 μm), for 5 min, CCh was applied (atropine+CCh). C, Dependence of DHPG-induced [Ca2+]i increases on intracellular Ca2+ stores. Time-lapse imaging of fluorescence changes (top) and voltage responses induced by APs at 30 Hz (bottom) were taken before (−DHPG) and during (+DHPG) bath application of 30 μm DHPG. An endoplasmic reticulum Ca2+/ATPase blocker, CPA, is known to cause depletion of the stores. After application of 20 μm CPA for 7.5 min, DHPG was applied (CPA+DHPG). D, Dependence of DHPG-induced [Ca2+]i increases on IP3 receptor activation. Fluorescence changes (top) were taken before (−DHPG) and during (+DHPG) bath application of 30 μm DHPG. Recordings were made with a pipette containing solution with an impermeable IP3 receptor blocker, 2 mg/ml heparin (right), or no additives (left). Voltage responses induced by APs at 30 Hz at the middle are shown on an expanded timescale at the bottom. The arrows indicate GPCR-mediated membrane depolarization during AP firing. A set of measurements was done using slices derived from an animal. The perfusate contained 20 μm CNQX and 50 μm APV in all of the experiments.