Administering high doses of tetanus toxin to animals produces neuromuscular blockade. Previous studies, in which specific F(ab) antibody fragments were used to mask the 50,000 MW COOH-terminal portion of the heavy chain (fragment c) on the toxin molecule, have shown that the paralyzing effect of the toxin was most probably located in an area comprising the light chain and the 50,000 MW NH2-terminal portion of the heavy chain (Fragment Ibc). In our study, the toxin was also complexed with F(ab) fragments directed to the light chain (alpha), heavy chain (beta), beta minus IIc, and with monoclonal antibodies to epitopes on IIc and beta minus IIc. Investigating the effect of the resulting complexes both in mice and on the sphincter pupillae muscle in rabbits permitted us to circumscribe further the tetanus toxin neuromuscular blocking activity in a region of the NH2-terminal fragment (Mr = 50,000) of the heavy chain (fragment beta minus IIc). Our results are consistent with the assumption that the beta minus IIc fragment is critical for the neuromuscular blockade activity of tetanus toxin. However, it cannot be ruled out that both the peripheral and central effects of the toxin result from the same portion of the toxin molecule, the nature of the action depending on where the toxin is carried after its introduction into the organism.

The effects of isotonic saline, cyclic AMP, cyclic GMP and their dibutyryl derivatives on normal and tetanus toxin-paralyzed sphincter pupillae muscles were studied. Isoproterenol was used to test the functional integrity of dilator muscles. All drugs, diluted either in isotonic saline or phosphate buffered saline, were injected into the anterior chambers of the eyes of New Zealand albino rabbit 72 hr after isotonic saline or 1000 minimum lethal doses (mouse MLD) of purified tetanus toxin. The results indicated a functional adrenergic receptor system in both saline and toxin-treated eyes. Cyclic nucleotides had no marked effect on the pupillary muscles in saline-treated, non-paralyzed rabbit eyes. Injections of cyclic AMP diluted in either isotonic saline or phosphate buffered saline had little or no effect on normal or paralyzed pupillary muscles. However, cyclic GMP injections temporarily reversed the paralysis of sphincter pupillae muscles. These observations provide further evidence that cyclic nucleotides are implicated in the mode of action of tetanus toxin.

Generalized nephrotoxicity was observed in the rat as early as 6 hr after intramuscular injection of 1000 mld of tetanus toxin. The most striking morphological changes, including swelling and rupture of the mitochondria, sloughing of the microvilli, and dissolution of cells, were found exclusively in the renal proximal tubule. Acute tubular necrosis with degeneration of the organelles and disruption of the cell membrane were also encountered throughout the entire nephron.

Tetanus toxin injected intramuscularly induced no significant changes in the levels of glycine, GABA, glutamate, glutamine or aspartate in extracts of spinal cord from rats killed at timed intervals during the development of local and generalized tetanus. The amino acid contents in the hemisegment (longitudinal one‐half) of the spinal cord (L 2 ‐L 6 ) on the injected side (left gastrocnemius muscle) did not differ significantly from the contents in the hemisegment of the spinal cord on the non‐injected side. Nor were there any consistent changes in the contents of the amino acids in either hemisegment of the spinal cord as the tetanic symptoms became progressively more severe. Hence, the amino acid pool in the spinal cord was relatively stable despite the metabolic changes known to occur in tetanus. Our observations are consistent with the view of J ohnston , D e G roat and CURTIS (1969) who suggested that if glycine were indeed a spinal inhibitory neurotransmitter released by interneurons affected by tetanus toxin, the toxin should interfere with the release of the amino acid rather than deplete the transmitter stores.