Purpose: Ethanol exposure has been used to demonstrate the increase of oxidative stress to a variety of tissues. We studied the effect of ethanol on the response of isolated strips of rat bladder to in vitro hypoxia in the absence of glucose (in vitro ischemia). Secondly, we determined if alpha-lipoic acid (LA) could alter the response to ethanol + in vitro ischemia.
Methods: Sixty-four rats were used for the these experiments. Each rat was anesthetized and its urinary bladder excised. The bladder body was cut into two longitudinal strips and each strip mounted in individual baths filled with oxygenated Tyrodes solution containing glucose at 37 degrees C. Ethanol (0.3%, 1%, or 3%) was placed in the first six baths (two strips at each concentration). The last two baths did not receive ethanol. Each strip was incubated for 1 h and then stimulated with field stimulation at 2, 8, and 32 Hz. Each strip was stimulated with 10 microM carbachol, washed three times with fresh oxygenated buffer and ethanol re-added to their respective baths. Each strip was then stimulated with 120 mM KCl and washed three times as before. Strips were then subjected to 1 h in vitro ischemia (incubation in the absence of glucose with Tyrode's equilibrated with nitrogen instead of oxygen). During the ischemic period, each strip was stimulated for 5 s every 10 min by 32 Hz FS to simulate hyperreflexia. At the end of the hour, the tissues were incubated for an additional hour in the presence of oxygen + glucose and subjected to a second series of stimulations as before. At all times, ethanol was maintained in baths 1-6. In set 2, 1% ethanol was added to the first six baths. LA was added to every other bath, and the experiments performed as mentioned earlier.
Results: (a) Ethanol at 0.3% or 1% had no effect on the contractile responses prior to exposure to in vitro ischemia; 3% was inhibitory. (b) In vitro ischemia mediated a significant decrease in the contractile responses to all forms of stimulation except for carbachol. (c) Ethanol mediated a dose-response enhancement of the contractile dysfunctions caused by in vitro ischemia. (d) LA completely reversed the effects of ethanol on contractile responses following in vitro ischemia except for carbachol.
Conclusions: The results demonstrate that direct exposure to ethanol significantly enhanced contractile dysfunctions mediated by in vitro ischemia followed by re-oxygenation and that the presence of LA significantly inhibits this effect of ethanol.