Isovolumetric regulation mechanisms in cultured cerebellar granule neurons

Cultured cerebellar granule neurons exposed to gradual reductions in osmolarity (- 1.8 mOsm/min) maintained constant volume up to - 50% external osmolarity (pi (o)), showing the occurrence of isovolumetric regulation (IVR). Amino acids, Cl-, and K+ contributed at different phases of IVR, with early eff lux threshold for [H-3]taurine, D-[H-3]aspartate (as marker for glutamate) of pi (o) - 2% and - 19%, respectively, and more delayed thresholds of - 30% for [H-3]glycine and - 25% and - 29%, respectively, for Cl- (I-125) and K+ (Rb-86). Taurine seems preferentially involved in IVR, showing the lowest threshold, the highest efflux rate (five-fold over other amino acids) and the largest cell content decrease. Taurine and Cl- efflux were abolished by niflumic acid and Rb-86 by 15 mm Ba2+. Niflumic acid essentially prevented IVR in all ranges of pi (o). Cl--free medium impaired IVR when pi (o) decreased to - 24% and Ba2+ blocked it only at a late phase of - 30% pi (o). These results indicate that in cerebellar granule neurons: (i) IVR is an active process of volume regulation accomplished by efflux of intracellular osmolytes; (ii) the volume regulation operating at small changes of pi (o) is fully accounted for by mechanisms sensitive to niflumic acid, with contributions of both Cl- and amino acids, particularly taurine; (iii) Cl- contribution to IVR is delayed with respect to other niflumic acid-sensitive osmolyte fluxes (osmolarity threshold of - 25% pi (o)); and (iv), K+ fluxes do not contribute to WR until a late phase (< - 30% Tio).