Role of Nitric Oxide in Neuromuscular Transmission and Its Effects at Different Frequencies of Nerve Stimulation

Background:The free radical gas nitric oxide (NO) exhibits diverse vital roles in the
human body.It is now recognized as a major messenger molecule. Neural NO-synthase is present in the sarcolemma of type II skeletal muscle fibers. In rats, the NO
synthase pathway is present in skeletal muscle, vascular smooth muscle and motor
nerve terminal. However, previous studies did not determine whether NO facilitates
or impairs neuromuscular transmission in preparations indirectly stimulated at
different frequencies. Aim of work: The study aims to examine the effect of NO in rat
neuromuscular preparation at different stimulation frequencies and modulation of its
effect by hemoglobin (NO scavenger). Methods: 30 rats were used in the experiment
and were divided into 2 groups: GpI: rat diaphragms were electrically stimulated by
supramaximal stimuli, at low frequency of 0.5Hz for 0.5msec, directly and indirectly
to induce simple muscle twitch, GpII: rat diaphragms were electrically stimulated by
high frequency of 100Hz, directly and indirectly to induce tetanic contraction. Rat
diaphragms were bathed in Krebs solution. To investigate the effect of NO, L-arginine was added to the bath in a dose of 4.7nM/50ml bath. Then bovine Hb (50
nM /50ml bath was added to scavenge NO. A contact time of 3 minutes is allowed for
each step and the amplitude of maximal contraction(∆Y), contraction time(∆X), and
1/2 relaxation time (1/2Rt) were measured in GpI, while only amplitude of maximal
contraction was measured in GpII. Results: NO significantly increased ∆Y, ∆X and
decreased 1/2 Rt when rat diaphragm preparations were stimulated indirectly at low
or high frequencies. In contrast, when rat diaphragm preparations were stimulated
directly at either low or high frequencies, NO significantly decreased ∆Y, ∆X, and
increased 1/2 Rt. Bovine Hb completely reversed the NO effects. Conclusion: We can
conclude that NO has dual actions, facilitatory and inhibitory, on skeletal muscle
contraction using indirect or direct electrical stimulation respectively at both low and
high frequencies. Bovine Hb antagonized the effects of NO in all experimental steps,
giving an additional proof that the recorded changes were NO mediated.