LOPERAMIDE BLOCKS HIGH-VOLTAGE-ACTIVATED CALCIUM CHANNELS AND N-METHYL-D-ASPARTATE-EVOKED RESPONSES IN RAT AND MOUSE CULTURED HIPPOCAMPAL PYRAMIDAL NEURONS

The effects of the antidiarrheal agent loperamide on high-voltage-acti vated (HVA) calcium channel activity and excitatory amino acid-evoked responses in two preparations of cultured hippocampal pyramidal neuron s were examined. In rat hippocampal neurons loaded with the calcium-se nsitive dye fura-2, rises in intracellular free calcium concentration ([Ca2+](i)) evoked by transient exposure to 50 mM K+-containing medium [high extracellular potassium concentration ([K+](o))] were mediated by Ca2+ flux largely through nifedipine-sensitive Ca2+ channels, with smaller contributions from omega-conotoxin GVIA (omega-CgTx)-sensitive Ca2+ channels and channels insensitive to both nifedipine and omega-C gTx. Loperamide reversibly blocked rises in [Ca2+](i) evoked by high [ K+](o) in a concentration-dependent manner, with an IC50 of 0.9 +/- 0. 2 mu M. At the highest concentration tested (50 mu M), loperamide elim inated rises in [Ca2+](i) evoked by high [K+](o), a result otherwise a chieved only in Ca2+-free medium or by the combined application of nif edipine, omega-CgTx, and funnel web spider venom to Ca2+-containing me dium. The action of loperamide was neither naloxone sensitive nor mimi cked by morphine and was seen at concentrations substantially less tha n those required to block influx of Ca2+ through the N-methyl-D-aspart ate (NMDA) receptor-operated ionophore. Similar results were obtained in cultured mouse hippocampal pyramidal neurons under whole-cell volta ge clamp. Voltage-activated Ca2+ channel currents carried by barium io ns (I-Ba) could be discriminated pharmacologically into nifedipine-sen sitive (L-type) and nifedipine-resistant, omega-CgTx-sensitive (N-type ) components. Loperamide (0.1-50 mu M) produced a concentration-depend ent reduction of the peak I-Ba with an IC50 value of 2.5 +/- 0.4 mu M and, at the highest concentration tested, could fully block I-Ba in th e absence of any other pharmacological agent. The loperamide-induced b lock was rapid in onset and offset, was fully reversible, and did not appear to be related to the known calmodulin antagonist actions of lop eramide. The current-voltage characteristics of the whole-cell I-Ba we re unaffected by loperamide and the block was not voltage dependent. L operamide also attenuated NMDA-evoked currents recorded at a membrane potential of -60 mV, with an IC50 of 73 +/- 7 mu M. The block of NMDA- evoked currents was not competitive in nature, was not reversed by ele vation of the extracellular glycine or spermine concentration, and was not affected by changes in the membrane holding potential. Steady sta te currents evoked by kainate and L-alpha-amino-3-hydroxy-5-methylisox azolepropionic acid were, in contrast, relatively unaffected by 100 mu M loperamide. We conclude that loperamide, applied at low micromolar concentrations, is a broad-spectrum blocker of neuronal HVA Ca2+ chann els. At higher concentrations, it reduces Ca2+ flux through NMDA recep tor-operated channels. Loperamide may prove to be a useful tool in exp eriments in which a general and reversible suppression of neuronal HVA Ca2+ channel activity is required.