Efficacy and Security of Tetrodotoxin in the Treatment of Cancer-Related Pain: Systematic Review and Meta-Analysis.

The pharmacological treatment of cancer-related pain is unsatisfactory. Tetrodotoxin (TTX) has shown analgesia in preclinical models and clinical trials, but its clinical efficacy and safety have not been quantified. For this reason, our aim was to perform a systematic review and meta-analysis of the clinical evidence that was available. A systematic literature search was conducted in four electronic databases (Medline, Web of Science, Scopus, and ClinicalTrials.gov) up to 1 March 2023 in order to identify published clinical studies evaluating the efficacy and security of TTX in patients with cancer-related pain, including chemotherapy-induced neuropathic pain. Five articles were selected, three of which were randomized controlled trials (RCTs). The number of responders to the primary outcome (≥30% improvement in the mean pain intensity) and those suffering adverse events in the intervention and placebo groups were used to calculate effect sizes using the log odds ratio. The meta-analysis showed that TTX significantly increased the number of responders (mean = 0.68; 95% CI: 0.19-1.16, p = 0.0065) and the number of patients suffering non-severe adverse events (mean = 1.13; 95% CI: 0.31-1.95, p = 0.0068). However, TTX did not increase the risk of suffering serious adverse events (mean = 0.75; 95% CI: -0.43-1.93, p = 0.2154). In conclusion, TTX showed robust analgesic efficacy but also increased the risk of suffering non-severe adverse events. These results should be confirmed in further clinical trials with higher numbers of patients.

Tetrodotoxin for Chemotherapy-Induced Neuropathic Pain: A Randomized, Double-Blind, Placebo-Controlled, Parallel-Dose Finding Trial.

Tetrodotoxin (TTX) has emerged as a potentially efficacious agent for chemotherapy-induced neuropathic pain (CINP), a prevalent, debilitating condition often resistant to analgesics. This randomized, double-blind, dose-finding study was undertaken to explore safety and trends in efficacy of four TTX doses and to identify a dose for further study. One hundred and twenty-five patients with taxane- or platinum-related CINP received subcutaneous placebo or TTX (7.5 µg twice daily (BID), 15 µg BID, 30 µg once daily (QD), 30 µg BID) for four consecutive days. Primary outcome measure was average patient-reported Numeric Pain Rating Scale (NPRS) score during Days 21-28 post-treatment. Changes in mean NPRS score were not statistically different between cohorts, due to small trial size and influence of a few robust placebo responders. Cumulative responder analysis showed significant difference from placebo with 30 µg BID cohort using the maximum response at any timepoint (p = 0.072), 5-day (p = 0.059), 10-day (p = 0.027), and 20-day (p = 0.071) rolling averages. In secondary quality of life (QOL) outcomes, 30 µg BID cohort also differed significantly from placebo in a number of SF-36 and CIPN20 subscales. Most adverse events (AE) were mild or moderate with oral paresthesia (29.6%) and oral hypoesthesia (24.8%) as most common.

Safety, Tolerability, Pharmacokinetics, and Concentration-QTc Analysis of Tetrodotoxin: A Randomized, Dose Escalation Study in Healthy Adults.

Tetrodotoxin (TTX) is a highly specific voltage-gated sodium channel (VGSC) blocker in clinical evaluation as a peripheral-acting analgesic for chronic pain. This study presents the first published results of the safety including cardiac liability of TTX at therapeutic-relevant concentrations in twenty-five healthy adults. Randomized, double-blind, placebo-, and positive- (moxifloxacin) controlled study evaluated single ascending doses of 15 µg, 30 µg, and 45 µg TTX over 3 periods with a 7-day washout between each period. Subcutaneous injections of TTX were readily absorbed, reaching maximum plasma concentration (Cmax) within 1.5 h. Both extent of exposure (AUC) and Cmax increased in proportion to dose. No QT prolongation was identified by concentration-QTc analysis and the upper bounds of the two-sided 90% confidence interval of predicted maximum baseline and placebo corrected QTcF (ΔΔQTcF) value did not exceed 10 ms for all tetrodotoxin doses, thereby meeting the criteria of a negative QT study. Safety assessments showed no clinically relevant changes with values similar between all groups and no subject withdrawing due to adverse events. Paresthesia, oral-paresthesia, headache, dizziness, nausea, and myalgia were the most common TEAEs (overall occurrence ≥5%) in the TTX treatment groups. TTX doses investigated in this study are safe, well-tolerated, and lack proarrhythmic proclivity.

The Incidence of Tetrodotoxin and Its Analogs in the Indian Ocean and the Red Sea.

Tetrodotoxin (TTX) is a potent marine neurotoxin with bacterial origin. To date, around 28 analogs of TTX are known, but only 12 were detected in marine organisms, namely TTX, 11-oxoTTX, 11-deoxyTTX, 11-norTTX-6(R)-ol, 11-norTTX-6(S)-ol, 4-epiTTX, 4,9-anhydroTTX, 5,6,11-trideoxyTTX, 4-CysTTX, 5-deoxyTTX, 5,11-dideoxyTTX, and 6,11-dideoxyTTX. TTX and its derivatives are involved in many cases of seafood poisoning in many parts of the world due to their occurrence in different marine species of human consumption such as fish, gastropods, and bivalves. Currently, this neurotoxin group is not monitored in many parts of the world including in the Indian Ocean area, even with reported outbreaks of seafood poisoning involving puffer fish, which is one of the principal TTX vectors know since Egyptian times. Thus, the main objective of this review was to assess the incidence of TTXs in seafood and associated seafood poisonings in the Indian Ocean and the Red Sea. Most reported data in this geographical area are associated with seafood poisoning caused by different species of puffer fish through the recognition of TTX poisoning symptoms and not by TTX detection techniques. This scenario shows the need of data regarding TTX prevalence, geographical distribution, and its vectors in this area to better assess human health risk and build effective monitoring programs to protect the health of consumers in Indian Ocean area.

Liquid Chromatography with a Fluorimetric Detection Method for Analysis of Paralytic Shellfish Toxins and Tetrodotoxin Based on a Porous Graphitic Carbon Column.

Paralytic shellfish toxins (PST) traditionally have been analyzed by liquid chromatography with either pre- or post-column derivatization and always with a silica-based stationary phase. This technique resulted in different methods that need more than one run to analyze the toxins. Furthermore, tetrodotoxin (TTX) was recently found in bivalves of northward locations in Europe due to climate change, so it is important to analyze it along with PST because their signs of toxicity are similar in the bioassay. The methods described here detail a new approach to eliminate different runs, by using a new porous graphitic carbon stationary phase. Firstly we describe the separation of 13 PST that belong to different groups, taking into account the side-chains of substituents, in one single run of less than 30 min with good reproducibility. The method was assayed in four shellfish matrices: mussel (Mytillus galloprovincialis), clam (Pecten maximus), scallop (Ruditapes decussatus) and oyster (Ostrea edulis). The results for all of the parameters studied are provided, and the detection limits for the majority of toxins were improved with regard to previous liquid chromatography methods: the lowest values were those for decarbamoyl-gonyautoxin 2 (dcGTX2) and gonyautoxin 2 (GTX2) in mussel (0.0001 mg saxitoxin (STX)·diHCl kg(-1) for each toxin), decarbamoyl-saxitoxin (dcSTX) in clam (0.0003 mg STX·diHCl kg(-1)), N-sulfocarbamoyl-gonyautoxins 2 and 3 (C1 and C2) in scallop (0.0001 mg STX·diHCl kg(-1) for each toxin) and dcSTX (0.0003 mg STX·diHCl kg(-1) ) in oyster; gonyautoxin 2 (GTX2) showed the highest limit of detection in oyster (0.0366 mg STX·diHCl kg(-1)). Secondly, we propose a modification of the method for the simultaneous analysis of PST and TTX, with some minor changes in the solvent gradient, although the detection limit for TTX does not allow its use nowadays for regulatory purposes.

Tetrodotoxin alleviates acute heroin withdrawal syndrome: a multicentre, randomized, double-blind, placebo-controlled study.

1. Tetrodotoxin (TTX) is a powerful sodium channel blocker extracted from the puffer fish. The efficacy and safety of TTX as monotherapy for the treatment of acute heroin withdrawal syndrome were evaluated in the present study. This 7-day, multicentre, randomized, double-blind, placebo-controlled study was carried out between December 2008 and October 2009. In total, 216 patients who met the Diagnostic and Statistical Manual of Mental Disorders IV diagnosis of heroin addiction were recruited. After providing written informed consent, subjects were randomly assigned to double-blind treatment in one of the following groups: 5 µg TTX group (group 1), 10 µg TTX group (group 2) or the placebo group (group 3). 2. Evidence suggests that both 5 and 10 µg TTX significantly reduced withdrawal symptoms by day 3 compared with placebo, and there was no significant difference in the incidence of adverse events in the three groups. 3. In conclusion, this clinical trial shows that TTX (5 and 10 µg given t.i.d.) is effective in alleviating opiate withdrawal symptoms with few side-effects.

Neuromuscular adaptations to respiratory muscle inactivity.

Cervical spinal cord injury results in significant functional impairment. It is important to understand the neuroplasticity in response to inactivity of respiratory muscles in order to prevent any associated effects that limit functional recovery. Recent studies have examined the mechanisms involved in inactivity-induced neuroplasticity of diaphragm motor units. Both spinal hemisection at C2 (C2HS) and tetrodotoxin (TTX)-induced phrenic nerve blockade result in diaphragm paralysis and inactivity of axon terminals. However, phrenic motoneurons are inactive with C2HS but remain active after TTX. Diaphragm muscle fibers ipsilateral to C2HS display minimal changes post-injury. Neuromuscular transmission is enhanced following C2HS but impaired following TTX. Synaptic vesicle pool size at diaphragm neuromuscular junctions increases after C2HS, but decreases after TTX. Thus, inactivity-induced neuromuscular plasticity reflects specific adaptations that depend on inactivity at the motoneuron rather than at axon terminals or muscle fibers. These results indicate that neuromuscular transmission and functional properties of diaphragm fibers can be maintained after spinal cord injury, providing a substrate for functional recovery and/or specific therapeutic approaches such as phrenic pacing.

Lidocaine, tetrodotoxin and their effect on consolidation of spatial memory / Lidocaína, tetrodotoxina y su efecto sobre la consolidación de la memoria espacial

This study was aimed at comparing the effect of unilateral hippocampal inactivation with tetrodotoxin (TTX) and lidocaine on spatial memory consolidation. Both drugs block voltage-dependent sodium channels. However, TTX and lidocaine differ in the duration of their effects, with maximum TTX effect between 30 min and 120 min, washing out in 24 hours. Lidocaine maximum effect occurs 20-30 minutes after administration. Our experimental subjects, twenty-four 3-month-old Wistar rats, were unilaterally implanted with stainless-steel cannulae aimed at the right dorsal hippocampus. Animals received four daily trials for 5 consecutive days. Control injections of 1 µl saline, or inactivating injections of 5ng of TTX in 1 µl saline or lidocaine (2%) in 1 µl were made through a guide cannula 1 minute after the last trial from day 1 to day 4. Results showed that the groups that received TTX or lidocaine did not differ but were impaired regarding controls, suggesting that short-term consolidation processes can account for the memory impairment observed here (AU)

El objetivo de este estudio ha sido comparar el efecto de la inactivación unilateral del hipocampo con tetrodotoxina (TTX) o lidocaína sobre la consolidación de la memoria espacial. Ambas drogas bloquean los canales de sodio dependientes de voltaje, pero difieren en la duración de sus efectos, mostrando la tetrodotoxina su máxima actividad entre 30 y 120 minutos, eliminándose completamente a las 24 horas. En cambio, el máximo efecto de la lidocaína acontece entre 20-30 minutos tras su administración. Nuestros sujetos experimentales fueron canulados unilateralmente sobre hipocampo dorsal. Los animales recibieron cuatro ensayos diarios durante 5 días consecutivos. Las inyecciones control de 1 µl de un compuesto salino, o inyecciones inactivadoras de 5ng de TTX en 1 µl de salino o lidocaína (2%) en 1 µl de salino, se aplicaron a través de una cánula guía 1 minuto después del último ensayo desde los días 1 al 4. Los resultados mostraron que los grupos que recibieron TTX o lidocaína no difieren entre ellos en sus efectos, pero en cambio resultaron perjudicados en comparación con el grupo control, lo que sugiere que los procesos de consolidación a corto plazo pueden explicar la alteración mnésica que se observa en este trabajo (AU)

Lidocaine, tetrodotoxin and their effect on consolidation of spatial memory.

This study was aimed at comparing the effect of unilateral hippocampal inactivation with tetrodotoxin (TTX) and lidocaine on spatial memory consolidation. Both drugs block voltage-dependent sodium channels. However, TTX and lidocaine differ in the duration of their effects, with maximum TTX effect between 30 min and 120 min, washing out in 24 hours. Lidocaine maximum effect occurs 20-30 minutes after administration. Our experimental subjects, twenty-four 3-month-old Wistar rats, were unilaterally implanted with stainless-steel cannulae aimed at the right dorsal hippocampus. Animals received four daily trials for 5 consecutive days. Control injections of 1 microl saline, or inactivating injections of 5 ng of TTX in 1 microl saline or lidocaine (2%) in 1 microl were made through a guide cannula 1 minute after the last trial from day 1 to day 4. Results showed that the groups that received TTX or lidocaine did not differ but were impaired regarding controls, suggesting that short-term consolidation processes can account for the memory impairment observed here.

Tetrodotoxin reduces cue-induced drug craving and anxiety in abstinent heroin addicts.

BACKGROUND: Tetrodotoxin (TTX) is a neurotoxin found in puffer fish and other marine animals. New clinical studies suggest that low-dose TTX can safely relieve severe, treatment-resistant cancer pain. The therapeutic potential of TTX in addiction is supported by studies in laboratory animals. The purpose of this double-blind, placebo-controlled study was to assess the effect of a single intramuscular dose of TTX on cue-induced craving and anxiety in abstinent heroin addicts. METHODS: Forty-five abstinent heroin addicts were randomly assigned to three treatment groups: placebo, 5 microg TTX, or 10 microg TTX. Participants were exposed to a neutral video or a heroin-related video. Craving, anxiety, blood pressure, and heart rate were measured pre- and post-exposure. RESULTS: Heroin-related cues increased both craving and anxiety and had no effect on blood pressure and heart rate. A single dose of TTX dose-dependently attenuated the increases in craving and anxiety while having no effect on blood pressure or heart rate. CONCLUSION: The results suggest that low-dose TTX is acutely effective in reducing cue-induced increases in heroin craving and associated anxiety.

An open-label, multi-dose efficacy and safety study of intramuscular tetrodotoxin in patients with severe cancer-related pain.

Cancer pain is a prevalent and serious public health issue, and more effective treatments are needed. This study evaluates the analgesic activity of tetrodotoxin, a highly selective sodium channel blocker, in cancer pain. A Phase IIa, open-label, multicenter, dose-escalation study of intramuscular tetrodotoxin was conducted in patients with severe, unrelieved cancer pain. The study design called for six ascending dose levels of intramuscular tetrodotoxin, administered over a four-day treatment period in hospitalized patients, with six patients to be enrolled within each successive dose level. Twenty-four patients underwent 31 courses of treatment at doses ranging from 15 to 90 microg daily, administered in divided doses, over four days. Most patients described transient perioral tingling or other mild sensory phenomena within about an hour of each treatment. Nausea and other toxicities were generally mild, but two patients experienced a serious adverse event, truncal and gait ataxia, that resolved over days. Seventeen of 31 treatments resulted in clinically meaningful reductions in pain intensity, and relief of pain persisted for up to two weeks or longer. Two patients had opioids held due to narcosis concurrent with relief of pain. Somatic, visceral, or neuropathic pain could all respond, but it was not possible to predict which patients were more likely to have an analgesic effect. Tetrodotoxin was overall safe. It effectively relieved severe, treatment-resistant cancer pain in the majority of patients and often for prolonged periods after treatment. It may have a novel mechanism of analgesic effect. Further study is warranted.

Reversible inactivation of amygdala and cerebellum but not perirhinal cortex impairs reactivated fear memories.

The cerebellum, amygdala and perirhinal cortex are involved in fear learning but the different roles that these three structures play in aversive learning are not well defined. Here we show that in adult rats amygdala or cerebellar vermis blockade causes amnesia when performed immediately, but not 1 h, after the recall of fear memories. Thus, the cerebellum, as well as the amygdala, influences long-term fear memories. These effects are long lasting, as they do not recover over time, even after a reminder shock administration. However, all of the subjects were able to form new fear memories in the absence of inactivation. By increasing the strength of conditioning, we observed that stronger fear memories are affected by the combined but not independent amygdala and cerebellar blockade. These results demonstrate that the cerebellum supports the memory processes even in the absence of a crucial site for emotions like the amygdala. Furthermore, they suggest that the amygdala is only one of the neural sites underlying long-term fear memories. Finally, the inactivation of the perirhinal cortex never alters retrieved fear traces, showing important differences between the amygdala, cerebellum and perirhinal cortex in emotional memories.

Beyond memory, navigation, and inhibition: behavioral evidence for hippocampus-dependent cognitive coordination in the rat.

Injecting tetrodotoxin (TTX) into one hippocampus impaired avoidance of a place defined by distal cues while rats were on a slowly rotating arena. The impairment could be explained by a deficit in memory, navigation, or behavioral inhibition. Here, we show that the TTX injection abolished the ability of rats to organize place-avoidance behavior specifically when distal room and local arena cues were continuously dissociated. The results provide evidence that injecting TTX into one hippocampus specifically impaired the coordination of representations that support organized behavior because of the following: (1) rats normally coordinate separate room and arena avoidance memories; (2) the TTX injection spared spatial, relational, and representational memory, navigation, and behavioral inhibition; and (3) the TTX-induced impairment of place avoidance depended on the need to coordinate representations of local and distal stimuli.

Myogenic constriction is increased in mesenteric resistance arteries from rats with chronic heart failure: instantaneous counteraction by acute AT1 receptor blockade.

(1) Increased vascular resistance in chronic heart failure (CHF) has been attributed to stimulated neurohumoral systems. However, local mechanisms may also importantly contribute to set arterial tone. Our aim, therefore, was to test whether pressure-induced myogenic constriction of resistance arteries in vitro--devoid of acute effects of circulating factors--is increased in CHF and to explore underlying mechanisms. (2) At 12 weeks after coronary ligation-induced myocardial infarction or SHAM-operations in rats, we studied isolated mesenteric arteries for myogenic constriction, determined as the active constriction (% of passive diameter) in response to stepwise increase in intraluminal pressure (20 - 160 mmHg), in the absence and presence of inhibitors of potentially involved modulators of myogenic constriction. (3) We found that myogenic constriction in mesenteric arteries from CHF rats was markedly increased compared to SHAM over the whole pressure range, the difference being most pronounced at 60 mmHg (24+/-2 versus 4+/-3%, respectively, P<0.001). (4) Both removal of the endothelium as well as inhibition of NO production (L-N(G)-monomethylarginine, 100 micro M) significantly increased myogenic constriction (+16 and +25%, respectively), the increase being similar in CHF- and SHAM-arteries (P=NS). Neither endothelin type A (ET(A))-receptor blockade (BQ123, 1 micro M) nor inhibition of perivascular (sympathetic) nerve conduction (tetrodotoxin, 100 nM) affected the myogenic response in either group. (5) Interestingly, increased myogenic constriction in CHF was fully reversed after angiotensin II type I (AT(1))-receptor blockade (candesartan, 100 nM; losartan, 10 micro M), which was without effect in SHAM. In contrast, neither angiotensin-converting enzyme (ACE) inhibition (lisinopril, 1 micro M; captopril, 10 micro M) or AT(2)-receptor blockade (PD123319, 1 micro M), nor inhibition of superoxide production (superoxide dismutase, 50 U ml(-1)), TXA(2)-receptor blockade (SQ29,548, 1 micro M) or inhibition of cyclooxygenase-derived prostaglandins (indomethacin, 10 micro M) affected myogenic constriction. (6) Sensitivity of mesenteric arteries to angiotensin II (10 nM - 100 micro M) was increased (P<0.05) in CHF (pD(2) 7.1+/-0.4) compared to SHAM (pD(2) 6.2+/-0.3), while the sensitivity to KCl and phenylephrine was not different. (7) Our results demonstrate increased myogenic constriction in small mesenteric arteries of rats with CHF, potentially making it an important target for therapy in counteracting increased vascular resistance in CHF. Our results further suggest active and instantaneous participation of AT(1)-receptors in increased myogenic constriction in CHF, involving increased sensitivity of AT(1)-receptors rather than apparent ACE-mediated local angiotensin II production.

Paralytic toxins in three new gastropod (Olividae) species implicated in food poisoning in southern Taiwan.

The toxins in the new gastropods Oliva miniacea, O. mustelina and O. nirasei implicated in a food paralytic poisoning incident in South Taiwan in February 2002 were studied. It was found that the three species of gastropods contained moderate amounts of toxin in edible portion only, and the highest toxicity score was 18 MU/g for O. miniacea, 10 MU/g for O. mustelina, and 27 MU/g for O. nirasei. The toxin was partially purified from the toxic specimens of each species by ultrafiltration using a YM-1 membrane, followed by chromatography on Bio-Gel P-2 column. Analyses by HPLC, GC-MS and LC-MS showed that the toxin from O. miniacea, O. nirasei and O. mustelina contained TTX, and related compounds 4-epi TTX and anhydro-TTX. The paralytic shellfish poisons were not found.

Identification of tetrodotoxin in marine gastropods implicated in food poisoning.

Attempts were made to identify the toxin in the gastropod Zeuxis samiplicutus that was responsible for the recent food poisoning incidents in southern Zhejiang, Mainland China. Symptoms associated with the poisonings included paralysis, coma, vomiting, and aphasia. The remaining specimens of gastropod were assayed for tetrodotoxin toxicity (TTX). The range of specimen toxicity was found to be 4 - 186 mouse units (MU), and the average toxicity was 111 +/- 45 MU. The toxin was partially purified from the methanol extract of the gastropod by ultrafitration and Bio-Gel P-2 column chromatography. Cellulose acetate membrane electrophoresis, TLC, and HPLC analyses demonstrated that the toxin contained TTX. It was concluded that the causative agent of the above food poisoning was TTX.

Toxicity and toxin profiles of the newt, Cynops pyrrhogaster from western Japan.

A total of 382 specimens of a Japanese newt, Cynops pyrrhogaster, were collected from western Japan during 1996 to 1999, and assayed for their individual, geographical, sexual, seasonal variations, and anatomical distribution of toxicity by mouse. Most of the specimens tested showed toxicity scores ranging from 5 to 370 MU/g, where no seasonal, but large individual, sexual, and regional variations of toxicity were clearly recognized. Among the parts, skin and muscle showed higher toxicity scores (56 MU/g) than liver, stomach, intestine and gonad, whose toxicity ranged from less than 2 to 33 MU/g. The C. pyrrhogaster toxin was purified by several steps of column chromatography and was shown to consist of tetrodotoxin (TTX) and 6-epiTTX as main components, and 4-epiTTX, 4,9-anhydro-6-epiTTX, and 4,9-anhydroTTX as minor ones by means of HPLC and 1H-NMR analyses.

Changes in electrophysiological properties of tibial motoneurones in the rat following 4 weeks of tetrodotoxin-induced paralysis.

In this study, we test the hypothesis that 4 weeks tetrodotoxin (TTX) paralysis altered the passive membrane properties of rat tibial motoneurones. Impulse activity along the sciatic nerve was blocked for 4 weeks using TTX delivered by an osmotic minipump to a Silastic cuff placed around the nerve. That portion of the sample exhibiting the 20% slowest After-hyperpolarization (AHP) decay time (AHPd), and which therefore included presumptive type S motoneurons, demonstrated responses (reduced AHPd, increased rheobase and rheobase voltage), which were not evident in the rest of the sample (presumptive fast motoneurons), in which an increased AHPd, in fact, was found. The results thus support the hypothesis that retrograde signals from inactive slow and fast muscle fibers have different effects on their innervating motoneurones.

Experimental use of tetrodotoxin for corneal pain after excimer laser keratectomy.

PURPOSE: To determine the duration of anesthesia, effect on corneal reepithelialization, and systemic toxicity of topical tetrodotoxin (TTX) administered after excimer laser keratectomy. METHODS: Two groups of six rabbits each underwent excimer laser keratectomy in the right eye to create a 5-mm-diameter wound, 75 mm in depth. One group then received a 40-microl aliquot of topical 1 mM TTX into the injured eye, whereas the other group received 40 microl of the sodium citrate vehicle as a control. The rabbits were treated with TTX or vehicle again at 6, 12, 18, and 24 h. Corneal sensation was tested at 3, 6, 9, 12, 15, 18, 21, 24, 30, 32, and 40 h. To determine whether TTX inhibited corneal reepithelialization, compared with vehicle-treated control eyes, the healing rate of the epithelial defect was measured. RESULTS: Administration of TTX every 6 h for 24 h produced nearly complete anesthesia for > or = 30 h. At 32 h, 8 h after the final application of TTX, there was still significant anesthesia of the TTX-treated corneas (p = 0.0325, Wilcoxon test). Normal corneal sensation in all TTX-treated animals returned at 40 h, or 16 h after the final dose. In contrast, vehicle-treated eyes all had normal sensation for nearly the entire duration of the experiment. At 40 h, the TTX-treated eyes had slightly larger defects than vehicle-treated eyes, 7.85+/-1.74 versus 4.54+/-1.31 mm2 (p < 0.025, t test). However, at 49 h and thereafter, both groups were equally healed (p > 0.05, t test). No systemic toxicity was observed in any of the rabbits. CONCLUSION: Topical TTX is a long-acting and nontoxic local anesthetic in a rabbit model of excimer laser keratectomy.