NEURONAL SWELLING AND SURFACE-AREA REGULATION - ELEVATED INTRACELLULAR CALCIUM IS NOT A REQUIREMENT

Neurons are mechanically robust. During prolonged swelling, molluscan neurons can triple their apparent membrane area. They gain surface are a and capacitance independent of extracellular Ca concentration ([Ca]( e)), but it is unknown if an increase in intracellular Ca concentratio n ([Ca](i)) is necessary. If Ca for stimulating exocytosis is unnecess ary, it is possible that swelling-induced membrane tension changes dir ectly trigger surface area readjustments. If, however; Ca-mediated but not tension-mediated membrane recruitment is responsible for surface area increases, swelling neurons should sustain elevated levels of [Ca ](i). The purpose of this investigation is to determine if the [Ca](i) in swelling neurons attains levels high enough to promote exocytosis and if any such increase is required. Lymnaea neurons were loaded with the Ca concentration indicator fura 2. Calibration was performed in s itu using 4-bromo-A-23187 and Ca-ethylene glycol-bis(beta-aminoethyl e ther)-N,N,N',N'-tetraacetic acid (EGTA), with free Ca concentration ra nging from 0 to 5 mu M. Swelling perturbations (medium osmolarity redu ced to 25% for 5 min) were done at either a standard [Ca](e) or very l ow [Ca](e) level (0.9 mM or 0.13 mu M, respectively). In neither case did the [Ca](i) increase to levels that drive exocytosis. We also moni tored osmomechanically driven membrane dynamics [swelling, then format ion and reversal of vacuole-like dilations (VLDs)] with the [Ca](i) cl amped below 40 nM via ,2-bis(2-aminophenoxy)ethane-N,N,N',N'-tetraacet ic acid (BAPTA). [Ca](i) did not change with swelling, and VLD behavio r was unaffected, consistent with tension-driven, [Ca](i)-independent surface area adjustments. In addition, neurons with [Ca](i) clamped at 0.1 mu M sia an ionophore could produce VLDs. We conclude that, under mechanical stress, neuronal membranes are compliant by virtue of surf ace area regulatory adjustments that operate independent of [Ca](i). T he findings support the hypothesis that plasma membrane area is regula ted in part by membrane tension.