Involvement of voltage- and ligand-gated Ca2+ channels in the neuroexcitatory and synergistic effects of putative uremic neurotoxins

Background. Renal failure has been viewed as a state of cellular calcium toxicity due to the retention of small fast-acting molecules. We have tested this hypothesis and identified potentially neuroexcitatory compounds among a number of putative uremic neurotoxins by examining the acute in vitro effects of these compounds on cultured central neurons. The in vitro neuroexcitatory and synergistic effects of guanidinosuccinate and spermine were also examined in vivo. Methods. The acute effects of 17 candidate uremic neurotoxins on murine spinal cord neurons in primary dissociated cell culture were investigated using the tight-seal whole-cell recording technique. The compounds studied comprised low-molecular-weight solutes like urea, indoles, guanidino compounds, polyamines, purines and phenoles, homocysteine, orotate, and myoinositol. Currents evoked by these compounds were further examined using various ligand- and voltage-gated ion channel blockers. The acute in vivo effects of guanidinosuccinate and spermine were behaviorally assessed following their injection in mice. Results. It was shown that 3-indoxyl sulfate, guanidinosuccinate, spermine, and phenol evoked significant whole-cell currents. Inward whole-cell current evoked by 3-indoxyl sulfate was not blocked by any of the applied ligand- or voltage-gated ion channel blockers, and the compound appeared to influence miscellaneous membrane ionic conductances, probably involving voltage-gated Ca2+ channels as well. Phenol-evoked outward whole-cell currents were at least partly due to the activation of voltage-gated K+ channels, but may also involve a variety of other ionic conductances. On the other hand, inward whole-cell currents evoked by guanidinosuccinate and spermine were shown to be due to specific interaction with voltage- and ligand-gated Ca2+ channels. Guanidinosuccinate-evoked current was caused by activation of N -methyl-d-aspartate (NMDA) receptor-associated ion channels. Low (mumol/L) concentrations of spermine potentiated guanidinosuccinate-evoked current through the action of spermine on the polyamine binding site of the NMDA receptor complex, whereas current evoked by high (mmol/L) concentrations of spermine alone involved direct activation of voltage-gated Ca2+ channels. Finally, intracerebroventricular administration of 0.25 mumol/L spermine potentiated clonic convulsions induced by guanidinosuccinate. These neuroexcitatory and synergistic effects of guanidinosuccinate and spermine could take place at pathophysiologic concentrations. Conclusion. The observed in vitro and in vivo effects of uremic retention solutes suggest that the identified compounds could play a significant role in uremic pathophysiology. Some of the compounds tested displayed in vitro and in vivo neuroexcitatory effects that were mediated by ligand- and voltage-gated Ca2+ channels. The findings suggest a mechanism for the involvement of calcium toxicity in the central nervous system complications in renal failure with particular reference to guanidinosuccinate and spermine.