N-tert-butyl-alpha-phenylnitrone: A free radical trap with unanticipated effects on diaphragm function

The spin trap N-tert-butyl-alpha-phenylnitrone (PEN) has a high avidity for free radical species and hence functions as an antioxidant in many biological systems. As such, we hypothesized that PEN would have powerful antioxidant effects on muscle function. We examined the effects of PEN on directly stimulated in vitro (37 degrees C) rat diaphragm. First, a dose-response curve for the effects of PEN on force frequency (n = 8) was established by comparing PEN-treated muscle strips (0.01-10 mM) with time- and stimulus-matched control strips. Second, the effect of 1.0 mM PEN on muscle endurance (n = 8) was established. Our findings were as follows. 1) Compared with baseline, peak twitch and low-frequency muscle tensions increased in a dose-dependent fashion, with peak effects at 1.0 mM PEN. 2) Muscle function at all stimulation frequencies was depressed at doses above 1.0 mM PEN. 3) Complete inhibition at 10 mM PEN was reversed with caffeine administration or washout. 4) During early fatigue, 1.0 mM PEN facilitated force. However endurance time decreased in the PEN-treated group. We conclude that PEN has direct reversible dose-dependent effects on diaphragm function. However, facilitation of low-frequency forces and the lack of fatigue-attenuating properties suggest that PEN has atypical antioxidant effects on muscle function.