New findings on the effects of the carbonic anhydrase inhibitor acetazolamide on transmitter release at the mouse neuromuscular junction

AYELÉN I. GROISMAN; OSVALDO D. UCHITEL

Huerta Grande, Córdoba

Congreso; XXVIII Congreso Anual de la Sociedad Argentina de Investigación en Neurociencias; 2013

Sociedad Argentina de Neurociencias

Acetazolamide (AZ) is known to inhibit the action of carbonic anhydrase (CA), an enzyme responsible for regulating the extra- and intracellular pH. This drug is used as an anticonvulsant and for treatment of episodic ataxia type-2 patients, but its mechanism of action is still unknown. Electrophysiology recordings done at the levator auris longus ex vivo muscles (NMJs) in bicarbonate solution showed a rise of spontaneous end plate potentials in muscles treated with AZ (100 uM), (3.1±0.2/s vs 1.92±0.06/s control), whereas quantal content measured at low high and burst stimulation showed no differences between treated and control NMJs. We further investigated the mechanism of action of AZ by studying the dynamics of vesicle exocytosis/endocytosis. We used fluorescence FM 2-10 dye in bicarbonate buffer to load (20Hz) and unload (50Hz) nerve terminals. The amount of dye loaded in the presence of AZ was reduced to 48±9% of control. After loading the nerve terminal in the absence of AZ followed by unloading the dye during 9 min in the presence of AZ, different kinetics were evidenced with a fluorescence retention (fr) of 68±5% in the treated NMJ in contrast to 8±4% in control. To gain further insight about the role of CA in the AZ mechanism of action unloading experiments were repeated in a Hepes (10mM, pH7.4) buffer. Experiments showed no effect of AZ in this condition:26±4% fr at treated NMJs vs 23±2% at control suggesting that AZ affects vesicle recycling through CA inhibition.