DIFFERENCES IN THE MULTIPLE STEP PROCESS OF INHIBITION OF NEUROTRANSMITTER RELEASE INDUCED BY TETANUS TOXIN AND BOTULINUM NEUROTOXINS TYPE-A AND TYPE-B AT APLYSIA SYNAPSES

In order to gain insights into the steps (binding, uptake, intracellul ar effect) which differ in the inhibitory actions of tetanus toxin and botulinum neurotoxins types A or B, their temperature dependencies we re investigated at identified cholinergic and non-cholinergic synapses in Aplysia. Upon lowering the temperature from 22 degrees C to 10 deg rees C, extracellularly applied botulinum neurotoxin type A and B appe ared unable to inhibit transmitter release whilst tetanus toxin exhibi ted a residual activity. Binding of each toxin to the neuronal membran e appeared virtually unaltered following this temperature change. By c ontrast, the intracellular effects of botulinum neurotoxin type B acid tetanus toxin were strongly attenuated by temperature reduction where as the inhibitory action of botulinum neurotoxin type A was only moder ately reduced. Importantly, this discrepancy relates to the known prot eolytic cleavage of different synaptic proteins by these two toxin gro ups. Since both the binding and intracellular activity of botulinum ne urotoxin type A are minimally affected at 10 degrees C, its inability to inhibit neurotransmission at this low temperature when applied extr acellularly indicated attenuation of its uptake. Due to the strict tem perature dependence of the intracellular action of tetanus toxin and b otulinum neurotoxin type B, but not A, an examination of the effects o f changes in temperature on the internalization step was facilitated b y the use of heterologous mixtures of the toxins' heavy and light chai ns. At 10 degrees C, heavy chain from tetanus toxin but not from botul inum neurotoxin type B mediated uptake of botulinum neurotoxin type A light chain. Collectively, these results provide evidence that, at lea st in Aplysia, the uptake mechanism for botulinum neurotoxin types A a nd B differs from that of tetanus toxin.