ATPase and carbonic anhydrase activities of bulk-isolated neuron, glia and synaptosome fractions from rat brain

Abstract

The (Na + + K +) ATPase, carbonic anhydrase and HCO 3 −-stimulated ATPase activities of bulk-isolated neuronal perikarya, glial and synaptosome fractions from 20–26-day-old rats were studied. The effects of varying K +, Na + and ATP concentrations were investigated to determine limits on how these enzymes might respond to changes in the levels of these substances in vivo. The (Na + + K +) ATPase activity of all three fractions had a similar high affinity for K +, with K m values in the range of 0.7–1.7mM. The K m for Na + was around 10-fold higher, in the range of 10–15mM. K m values for ATP were also not markedly different between the different fractions, being 1.2mM for the neurons and 1.8mM for the glia and synaptosome fractions, respectively. The V max at infinite [ATP] in the presence of 10mM K + was 2.9-fold higher for the glia as compared to the neuron fraction and 1.5-fold higher than the synaptosome fraction. At infinite K + in the presence of 3.3mM ATP the corresponding figures were 3.0 and 2.1. Arrhenius plots of (Na + + K +) ATPase activity were different for the neuron and glia fractions as compared to the synaptosome fraction,, suggesting subtle differences in themembrane environment and/or the enzyme molecule itself. The HCO 3 −-stimulated ATPase activity was only 14% higher in the glial fraction compared to the neurons and was variably stimulated by added K + at concentrations < 10mM. The glial-enriched cell fraction had a 2-fold higher specific carbonic anhydrase activity than the neuron fraction, but the combined activity of these fractions only represented < 2% of the total brain activity. Total brain carbonic anhydrase activity was not stimulated by added K + in the range of 5–50mM K +. These data indicate that none of the enzymes studied are likely to directly respond to extracellular K + levels in excess of 10mM, and therefore would only be secondarily involved in cerebrocortical swelling caused by increasing K + concentrations 20mM. The relation of these findings to previous work on increased (Na + + K +) ATPase and carbonic anhydrase activity in glial cells, the role of glial cells in passive or mediated transport processes related to increased levels of extracellular K + and the suitability of bulk-isolated glia and neuron-enriched fractions as experimental models are discussed.