Effects of TMB-8, a putative calcium antagonist, on neuromuscular transmission and muscle contractility in the mouse phrenic nerve-hemidiaphragm preparation

The effects of TMB-8 [8-(N.N.-diethylamino)octyl-3,4,5-trimethoxybenzoate], a putative calcium antagonist, on directly and indirectly evoked isometric twitches, tetanic contractions and potassium- and caffeine-induced contractures, were investigated in the mouse isolated phrenic nerve-hemidiaphragm preparation. In the lowest concentration tested (10^-6 M), TMB-8 produced an augmentation of both directly and indirectly induced twitches. In higher concentrations (10^-5-3 x 10^-5 M), this augmentation was followed by twitch reduction. In the highest concentrations (10^-4 M-3 x 10^-4 M), only twitch reduction in a concentration-dependent manner was observed. TMB-8 also depressed both directly and indirectly induced tetanic contractions. However, the drug was more effective in depressing neurotransmission than in reducing muscle contractility. Elevated Ca²⁺ (4-8 mM) or 3,4-diaminopyridine (10^-4 M) produced a good reversal of neuromuscular blockade but this effect was transient. Pretreatment with 4 mM Ca²⁺ had no significant effect on the time required to produce a 50% or a 90% inhibition of directly or indirectly induced twitches. However, 8 mM Ca²⁺ significantly prolonged the inhibitory effects of TMB-8 on indirectly, but not directly induced twitches. On the other hand, neostigmine (3 μM) appeared to hasten the blockade of transmission. Submaximal potassium-induced contractures were markedly depressed while caffeine-induced contractures were only slightly depressed by TMB-8 in the concentration range tested (10^-5-3 x 10^-4 M). The results are consistent with the hypothesis that TMB-8 inhibits skeletal muscle contractility by a reduction in transmembrane Ca²⁺ movement, a depression of postsynaptic acetylcholine receptor sensitivity, and a decreased mobilization of sequestered calcium from the sarcoplasmic reticulum.