LACTATE AND PYRUVATE LEVELS IN THE MOUSE OVIDUCT

The availability of lactate and pyruvate, required for in vitro mouse embryo development, in the embryo's natural environment was determined by measuring their levels in oviductal tissues. Factors controlling these levels were investigated by comparing oviducts from cycling (non-mated), pregnant and pseudopregnant animals during the first five days following ovulation.;During the normal estrus cycle, pyruvate levels in the ampulla and isthmus showed a moderate increase, measured 3 hours after ovulation (2.6 mmoles(.)kg('-1)) as compared with 12 hours before ovulation (1.8 mmoles(.)kg('-1)), while lactate levels were not increased until 12 to 24 hours after ovulation (24-28 mmoles(.)kg('-1) vs 16-19 mmoles(.)kg('-1) before ovulation). . . . (Author's abstract exceeds stipulated maximum length. Discontinued here with permission of school.) UMI;Quantitative, sensitive, and specific histochemical assays were developed for the measurement of pyruvate and lactate in nanogram-sized samples of freeze-dried tissue, using 'oil-well' techniques. The assays utilized lactate dehydrogenase (LDH) for the reduction of pyruvate or the oxidation of lactate. The pyridine nucleotide product was analyzed via emzymatic cycling, with a fluorometric assay of malate, the final product of the cycling reaction. The method for measurement of each metabolite was tested to insure a quantitative, sensitive, and specific assay in the tissues used. Experiments were conducted to determine the method of collecting tissues, which best maintained in vivo metabolite levels. Decapitation, rapid dissection of the oviducts from the body, and quick freezing was accomplished in an average of 15 seconds. With extended ischemia (45 seconds) beyond the normal collection time, these metabolites accumulated relatively slowly with pyruvate increasing 37% and lactate 32%. Pentobarbital anesthesia, with or without a 15 second period of anoxia (interruption of blood flow) produced a 54% decrease in pyruvate compared to unanesthetized controls, but had no effect on lactate concentration. The use of anesthesia in the collection procedure was rejected because of this apparent inhibition of metabolism.