In vivo fluorometric measurement of changes in cytosolic free calcium from the cat cortex during anoxia.

Abstract

A new approach to assess the mean changes in intracellular free calcium [Ca2+]i directly from the cortex in situ is described along with the [Ca2+]i changes during nitrogen anoxia. Following incision of the dura and part of the pia-arachnoid membrane, quin2 acetoxymethyl ester, 100 microM in artificial CSF, was superfused for 60 min onto the cat cortex. A small cortical area was irradiated with ultraviolet rays (350/30 nm) and the changes in the fluorescence and reflectance were recorded microfluorometrically at 506 and 366 nm, respectively. The net change in the quin2-Ca2+ fluorescence was calculated after correction for the hemodynamic artifact and subtraction of the basal NADH change. The quin2-Ca2+ fluorescence began to increase significantly (48.0 +/- 13.4 units; p less than 0.05) 20 s prior to the isoelectric electrocorticogram (ECoG) and remained elevated during nitrogen anoxia. It decreased steeply 7.3 +/- 1.7 s prior to the recovery of the ECoG activity after the animal was reoxygenated. Thus, the changes in the intracellular free calcium preceded those of the ECoG during a reversible anoxic insult, suggesting that the increase in the [Ca2+]i might be related to the electrical failure during anoxia.