Qualitative measurements of cytosolic calcium ion concentration within isolated guinea pig nerve endings using entrapped arsenazo III.
If arsenazo III is present during homogenization of brain this metallochromic indicator is entrapped within subsequently isolated synaptosomes. A large proportion of the entrapped indicator is released upon addition of digitonin to disrupt the synaptosomal plasma membrane. A similar proportion of [3H]sucrose is also trapped within synaptosomes if present in the homogenization medium, suggesting that homogenization causes a transient opening of the nerve ending as it is chopped off from the axon. Addition of the ionophore A23187 or depolarization of the plasma membrane by adding veratridine, gramicidin or increasing external K+ changes the absorbance of the entrapped dye, with peaks of absorbance around 600 and 650 nm, typical of the arsenazo III-Ca2+ complex. The response to veratridine is inhibited by the Ca2+-channel antagonist, verapamil, while that of A23187 is unaffected. The present method provides a sensitive technique for measurements of changes in cytosolic calcium ion concentrations within nerve endings.
D000818:Animals; D001150:Arsenazo III; D001391:Azo Compounds; D001693:Biological Transport, Active / Q000187:drug effects; D000001:Calcimycin / Q000494:pharmacology; D002118:Calcium / Q000378:metabolism*; D002462:Cell Membrane / Q000378:metabolism; D003600:Cytosol / Q000378:metabolism; D004533:Egtazic Acid / Q000494:pharmacology; D006168:Guinea Pigs; D007700:Kinetics; D008564:Membrane Potentials / Q000187:drug effects; D013395:Sucrose / Q000378:metabolism; D013574:Synaptosomes / Q000378:metabolism*; D014701:Veratridine / Q000494:pharmacology
D001391:Azo Compounds; D001150:Arsenazo III; D000001:Calcimycin; D004533:Egtazic Acid; D013395:Sucrose; D014701:Veratridine; D002118:Calcium