<p>Strength-duration (S-D) relationship reflects muscle excitability which partially determines force production. Isolated muscle contains part of nerve and neuromuscular junction, suggesting that muscle excitation initiates not only muscle membrane but also nerve terminals and hence neuromuscular transmission (NMT). This study aimed to examine the effects of inhibition of neuromuscular transmission on S-D relationship in slow- and fast-twitch muscles of mice. Isolated slow-twitch soleus (SOL) and fast-twitch extensor digitorum longus (EDL) muscles were electrically stimulated, and minimal field strength required for force generation [threshold field strength (Th_FS)] was determined at various duration. Th_FS significantly increased in the presence of NMT inhibitors including pancuronium (Pan) and succinylcholine in both SOL and EDL muscles. Moreover, Th_FS in the presence of NMT inhibitors was higher in SOL compared to EDL. To clarify the mechanism of the different Th_FS between SOL and EDL, contribution of Na⁺&#xa0;channel was investigated. In the presence of 1&#xa0;μM Pan, partial inhibition of Na⁺ channel by tetrodotoxin eliminated the difference in Th_FS between EDL and SOL. These results suggest that (1) NMT significantly enhances muscle excitability, (2) threshold of action potential in fast-twitch muscle is greater compared with slow-twitch muscle if no existence of NMT, and (3) Na⁺ channel likely contribute to the difference between slow- and fast-twitch muscles.</p>