Structure and pharmacokinetics
TTX is a small molecule (~319 Da) that features a guandininum moiety. This guanindinium is vital for TTX’s function as it essentially mimics a sodium ion to enter the VGSC. Based on the lethal dose of TTX in mice (LD = 334 μg/kg), about 25 mg of TTX is expected to kill a 75 kg human. Peak serum concentration of TTX occurs ~4 hours after ingestion, and it is excreted renally over the course of 2 – 4 days. The dissociation constant of the toxin varies depending on the tissue, but it ranges from 1 – 10 nM.
Models of TTX binding
Despite the evidence of many interactions, no actual widely accepted model of TTX binding to a sodium channel exists. Various diagrams showcasing the different researched interactions have been created with computer software, and even a molecular dynamics simulator. In one model, the guanidinium of TTX is shown interacting with Asp400 and Glu755 of DEKA while the C11 hydroxyl forms a hydrogen bond with another Glu (Figure 1). Unlike the cation-π interaction described in another study, this model shows Tyr401 forming a hydrophobic interaction with C4 – C8 segment of TTX. The researchers offered an explanation for this discrepancy: perhaps the cation-π interaction is only an intermediate one that allows TTX to enter the selectivity filter. The final confirmation however, involves the guanidinium interacting with DEKA.
The earliest and most common symptom is paresthesia, or a tingling sensation, of the lips and mouth and then eventually the entire body. This agrees with TTX’s action because the root cause of paresthesia is abnormal nerve impulses. TTX also acts on skeletal muscle, which eventually results in motor paralysis. The leading cause of death from TTX poisoning is respiratory failure. The exact cause of this failure is not phrenic nerve inactivation, but instead the cessation of diaphragm contraction. When mice are injected with TTX and respiration stops after 20 seconds but phrenic nerve impulses can be detected for up to 31 seconds, indicating that TTX has a greater effect on the smooth muscle of diaphragm, than on the phrenic nerve or medulla. As a result, respiratory and ventilatory support is mandated for patients who present with TTX poisoning symptoms, in some hospitals. One case study reported that a patient with complete respiratory failure was saved with immediate intubation and assisted ventilation.
Cardiac arrest can occur with TTX poisoning, but only if a very high dose was consumed. There are nine subtypes of VGSCs (NaV1.1 – 1.9), with channels Nav1.5, Nav1.8, and Nav1.9 being more resistant to TTX. Nav1.5 channels are primarily expressed in the heart, making the heart more resistant to TTX. The IC50 of TTX in cardiac cells is 100-fold higher than in nerve and skeletal muscle cells (1 μM vs. 10 nM). Nav1.5 channels are more resistant to TTX due to a Cys374 residue instead of a Tyr374 in the selectivity filter. Mutation of Cys to Tyr increases TTX affinity, and the reverse mutation decreases it. Researchers have postulated two models for this observation (Figure 2). In the first, Tyr enables a cation-π interaction with the guanidinium, stabilizing TTX. This interaction would not be possible with a Cys. The second model suggested that Cys forms a disulfide bond with another Cys, which would act as a physical barrier for TTX because it cannot displace the disulfide bond.