REGULATION OF CYTOSOL NUCLEUS PH GRADIENTS BY K+ H+ EXCHANGE MECHANISM IN THE NUCLEAR-ENVELOPE OF NEONATAL RAT ASTROCYTES/

In order to study the subcellular heterogeneity of intracellular H+ co ncentration in reactive astrocytes, the pH in the nucleus and cytosol of cultured astrocytes was measured using a confocal laser scanning mi croscope (CLSM) and pH indicator dye, 5'(and 6')-carboxyseminaphthoflu orescein (carboxy SNAFL-1). The change in intracellular pH was indexed by the fluorescence ratio( F-535/F-610) at an excitation wavelength o f 514.5 nm. The in vitro fluorescence ratio increased as pH decreased. This ratio in the nucleus was significantly lower than that in the cy tosol of astrocytes when perfused by HEPES-buffered Hanks' balanced sa lt solution (HHBSS) at pH 7.4. Acid stimulations of cells (pH 5.0) rai sed the fluorescence ratio in both nucleus and cytosol. However, the i ncrease in the fluorescence ratio of the nucleus was less than that of cytosol. Treatment with a K+/H+ ionophore, nigericin (20 mu M), rever sibly nullified this cytosol-nucleus pH gradient. These findings sugge st that a buffering mechanism(s) for maintaining of intracellular pH e xists between the nucleus and cytosol, and a K+/H+ exchanger may act o n the nuclear envelope to eventuate intranuclear pH maintenance in the living cells. (C) 1998 Elsevier Science B.V. All rights reserved.