Levels of adenosine and adenine nucleotides in slices of rat hippocampus

Abstract

ATP, ADP, AMP, IMP, adenosine, inosine and cyclic AMP were measured in slices of the rat hippocampus maintained in vitro. Immediately following cutting ATP was low (3.5 ± 0.6nmol/mg protein) and AMP high (8.6 ± 0.9nmol/mg), giving an energy charge ATP+12ADP)/(ATP + ADP + AMP). 0.34 ± 0.02. Over the next 90 min the energy charge gradually normalized (to 0.92 ± 0.01), partly due to conversion of AMP to ATP, but mainly to breakdown to adenosine and other purines which were recovered in the incubation medium. Total purine content decreased from approximately 18 to 10 nmol/mg protein in the first hour following cutting. In slices from old rats the energy charge was lower 60 min following preparation than in younger rats, while cyclic AMP and adenosine levels were higher. The adenosine antagonist 8-phenyl-theophylline tended to enhance the recovery of responsiveness after preparation of the slices. Stimulation of excitatory afferent fibers at a frequency of 10 Hz for 5 min did not significantly alter the purine levels in brain slices, while hypoxia decreased the energy charge significantly and tended to increase adenosine levels. These changes occurred somewhat later than the fall in electrophysiological responsiveness. 8-Phenyl-theophylline was able to delay somewhat the decline in the amplitude of synaptic responses under hypoxic conditions. It is concluded that the viable part of the hippocampal slice, which accounts for about half of the tissue, has levels of adenine nucleotides and adenosine which are similar to those found in the intact rat brain. The return of electrophysiological function following slice preparation is paralleled by a normalization of the energy charge, the adenosine level and the concentration of cyclic AMP. The absence of electrophysiological activity following cutting, and the decreases in such responses following either prolonged afferent stimulation or hypoxia may be related to changes in purine concentration in the slice. Although adenosine accumulating in the slice may contribute to the depression of electrophysiological responses it is probably not the major factor responsible for the reduction in synaptic responsiveness.