Summary
(1) Human platelets were incubated with [14C]adenine in plasma, washed and resuspended in salt solutions. The effects of incubating
the cells with antimycin A and 2- deoxyglucose on the radioactivity of ATP, ADP, AMP,
IMP and inosine + hypoxanthine, the total level of ADP and ATP and lactate production
were studied. The metabolic inhibitors only affected the cytoplasmic, [14C]-labelled nucleotides, and were apparently without effect on the granular, non-labelled
ATP and ADP. (2) Antimycin A caused a rapid, shortlasting decrease in [14C]ATP which was independent on glucose and enhanced by 2- deoxyglucose. Lactate production
increased about 3-fold with and without 2-deoxyglucose. The initial fall in [14C]ATP was therefore thought to be due to a failure to immediately substitute for the
lost oxidative ATP production. (3) After the initial fall in [14C]ATP no changes took place when glucose was present, while [14C]ATP and the adenylate energy charge decreased when glucose was absent and more so
when 2-deoxyglucose was present. Thus, glycogenolysis, corresponding to an ATP turnover
of 29.8 nmoles × min-1 × mg-1 protein did not maintain ATP homeostasis in platelets. (4) The changes in [14C]ATP, adenylate energy charge and lactate production under the various conditions
strongly suggested that regulation of ATP consumption in the cells was a major mechanism
to maintain ATP homeostasis. (5) The changes in the [14C]nucleotides in washed platelets were qualitatively similar to those previously described
in platelet-rich plasma (Holmsen et al. 1974); quantitative differences were apparent
and found to be due to binding of antimycin A to plasma proteins and counteraction
of the effects of 2-deoxyglucose by plasma glucose. In particular, the rapid, initial
fall in [14C]ATP occurred only with washed cells and clearly demonstrated a conversion of ATP
to hypoxanthine ADP, AMP, IMP and inosine. (6) The cellular AMP deaminase reaction
was found not to be related by the adenylate energy charge, in contrast to studies
with semipurified enzyme (Chapman & Atkinson 1973). Our results suggested that AMP
deamination occurred when [AMP] rose from its resting value of 0.07 mM and was inhibited
when [ATP] fell below 2 mM within the cell.
