• S J Dodgson and K Cherian.
• Department of Physiology, School of Medicine, University of Pennsylvania, Philadelphia 19104.
• Am. J. Physiol. 1989 Dec 1; 257 (6 Pt 1): E791-6.

AbstractAt 37 degrees C, pH 7.4, carbonic anhydrase activity (kenz) of disrupted rat renal proximal tubules and cortical mitochondria was 2.5 +/- 0.8 (n = 3) and 0.15 +/- 0.40 (n = 3) ml.mg-1.s-1, respectively. Turnover number for renal mitochondrial carbonic anhydrase (CA V) was 24,000 s-1. CA V activity of intact mitochondria was completely inhibited by 0.15 microM ethoxzolamide (EZ). Intact proximal tubules, prepared from 48-h starved male rats, were incubated at 37 degrees C in 10 mM pyruvate in Krebs-Henseleit bicarbonate saline buffer, 5% CO2-95% O2. The rate of glucose synthesis over 60 min was reduced 50% by including 0.6 microM EZ in the incubation solution. The concentration of NaHCO3 was doubled to 50 mM (with a corresponding decrease in NaCl) and the solution gassed with 10% CO2-90% O2; 2.4 microM EZ no longer decreased glucose synthesis. It was concluded that inhibition of glucose synthesis by EZ was directly a result of inhibiting the carbonic anhydrases. The rate of glucose production was subsequently determined with tubules incubating in a HCO3(-)-free N-2-hydroxyethylpiperazine-N'-2-ethane-sulfonic acid (HEPES) buffer; this rate was decreased 50% by 0.6 microM EZ. These data support the hypotheses that CA V provides HCO3- for pyruvate carboxylase and that CO2 can be provided by tubular metabolism. Intact tubules were incubated in from 5 to 20 mM pyruvate in either 25 or 50 mM HCO3-; in either buffer, the rate of glucose synthesis was similar, increasing with increasing pyruvate concentration. At no pyruvate concentration was there a change in the rate of glucose production when tubules were incubated in 50 mM HCO3- buffer with 1.6 microM EZ. These data also support the hypothesis that CA V provides the HCO3- substrate for pyruvate carboxylation when there is a high rate of intracellular CO2 production and external CO2 is low. It is further concluded that the cytosolic carbonic anhydrase (CA II) and the membrane-bound carbonic anhydrase (CA IV) are not involved in glucose synthesis from pyruvate.

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