Acetazolamide induces a unique and characteristic limb deformity in the offspring of pregnant rodents. The teratogenic mechanism of action is thought to be inhibition of carbonic anhydrase. One after effect of carbonic anhydrase inhibition is raised arterial and tissue carbon dioxide content and we have now demonstrated that maternal carbon dioxide inhalation leads to the same unique forelimb deformity substantiating the idea that carbonic anhydrase inhibition is the mechanism of acetazolamide teratogenesis. This application describes experiments which will attempt to uncover the mechanism by which C02 perturbs development. We hypothesize that C02 lowers intracellular pH (pHi) which subsequently leads to decreased proliferative rate thereby leading to the limb reduction deformity. We will measure pHi in the developing limb bud after teratogen treatment and attempt to correlate any changes with teratologic outcome and proliferative rate. The ability of limb bud cells to respond to internal pH changes will be documented and manipulation of this process will be correlated with teratologic outcome. We believe that reducing the intracellular pH of embryonic cells may be a general mechanism of teratogenesis and intend to study one human teratogen, alcohol, which could conceivably act through this mechanism.
