Cocaine Lethality Research Articles

Subunit Stabilization and Polyethylene Glycolation of Cocaine Esterase Improves In Vivo Residence Time

No small-molecule therapeutic is available to treat cocaine addiction, but enzyme-based therapy to accelerate cocaine hydrolysis in serum has gained momentum. Bacterial cocaine esterase (CocE) is the fastest known native enzyme that hydrolyzes cocaine. However, its lability at 37°C has limited its therapeutic potential. Cross-linking subunits through disulfide bridging is commonly used to stabilize multimeric enzymes. Herein we use structural methods to guide the introduction of two cysteine residues within dimer interface of CocE to facilitate intermolecular disulfide bond formation. The disulfide-crosslinked enzyme displays improved thermostability, particularly when combined with previously described mutations that enhance stability (T172R-G173Q). The newly modified enzyme yielded an extremely stable form of CocE (CCRQ-CocE) that retained greater than 90% of its activity after 41 days at 37°C, representing an improvement of more than 4700-fold over the wild-type enzyme. CCRQ-CocE could also be modified by polyethylene glycol (PEG) polymers, which improved its in vivo residence time from 24 to 72 h, as measured by a cocaine lethality assay, by self-administration in rodents, and by measurement of inhibition of cocaine-induced cardiovascular effects in rhesus monkeys. PEG-CCRQ elicited negligible immune response in rodents. Subunit stabilization and PEGylation has thus produced a potential protein therapeutic with markedly higher stability both in vitro and in vivo.

Capsaicin, an active ingredient in pepper sprays, increases the lethality of cocaine

Since 1992, California police have been using pepper sprays containing oleoresin capsicum (OC) as a nonlethal method to subdue delirious or violent individuals. Capsaicin is a primary ingredient in OC spray. From January 1993 to June 1995, at least 20 deaths in California were associated with OC and stimulant drug (cocaine, amphetamines, or ephedrines) exposure. Based on this background, we hypothesized a direct potentiation of cocaine toxicity by capsaicin. We performed animal experiments and also reviewed human data involving capsaicin and stimulants. The lethal effects of capsaicin administered with cocaine (both compounds administered intraperitoneally) were assessed in 14 groups of 20–40 male mice. Capsaicin at 10 mg/kg increased the lethality of cocaine in mice dosed at 60 mg/kg from 13% to 53% (P < 0.01) and for cocaine at 75 mg/kg from 53% to 90% (P < 0.001). Data from 26 autopsy and police reports from incidents in which death occurred after OC exposure were retrospectively analyzed for demographics, ethnicity, underlying disease, cause of death, and toxicology. All human deaths occurred in delirious and combative males, and 79% of these deaths occurred 1 h or less after OC exposure. Data indicated the presence of stimulants including cocaine in as many as 19 out of 26 deaths: 9 involved methamphetamine, 6 cocaine, 3 methamphetamine plus cocaine, and 1 pseudoephedrine. The animal experiments together with the human retrospective analysis support the idea that exposure to OC spray in cocaine-intoxicated individuals potentiates cocaine lethality.

Ecgonine methyl ester protects against cocaine lethality in mice.

Plasma cholinesterase (PChE) metabolizes cocaine to ecgonine methyl ester (EME). Limited data demonstrate that EME is a mild vasodilator. Exogenous PChE protects against cocaine-induced seizures and lethality. It is unclear whether this protective effect results from enhanced degradation of cocaine, the loss of active metabolites (benzoylecgonine, norcocaine), or the production of a beneficial metabolite (EME). This study was designed to further investigate the pharmacologic effects of EME. All experiments used female ICR Swiss albino mice weighing 20-30 grams. Mice were acclimated to 12 h alternating light-dark cycles and given food and water ad libitum. Using a randomized, blinded protocol, 80 mice were then pretreated with either IP EME (50 mg/kg) in a 0.9% sodium chloride solution or an equal volume of 0.9% sodium chloride solution as control. Five minutes later, all animals received 126 mg/kg of cocaine IP and were observed for seizures and death. Fatality was compared using a Fisher's exact test, and the time to seizures and death were compared using a Mann-Whitney U statistic. Pretreatment with EME increased survival following cocaine (9/40 vs. 2/40, for EME vs. control, respectively, p<0.05). The median times to seizure and death for both groups were 2.0 vs. 1.5 min (p>0.05), and 4.5 vs. 4.6 min (p>0.05) (EME vs. control for seizures and death, respectively). In this animal model, EME is protective against cocaine lethality. This effect is consistent with the previously described vasodilatory effects of EME. Further studies are indicated to determine whether the increase in EME produced by exogenous PChE administration contributes to the benefits that occur when PChE is given to cocaine-poisoned animals.

Effects of alcohol on cocaine lethality in rats: acute and chronic assessments

Cocaine and alcohol is a popular, yet toxic, drug combination that results in effects greater than that of either drug alone. The following experiment presents additional evidence that supports this position. Specifically, the lethal effects of acute (1 day) and chronic (4 days) alcohol (0.5 g/kg), cocaine (20, 30, or 40 mg/kg), or the respective alcohol/cocaine combinations were assessed in rats. For acute drug administration, lethality was only evident in those animals administered the combination of 0.5 g/kg alcohol and 40 mg/kg cocaine, supporting the position that the effects of combining alcohol and cocaine are greater than either drug alone. Chronic drug administration resulted in a weak sensitization to cocaine-induced lethality that was increased when alcohol was combined with cocaine. Together, this evidence suggests that combining alcohol and cocaine (acutely) can have lethal consequences in rats that are greater than either drug alone, effects that can be exacerbated with chronic use.

Capsaicin increases the lethality of cocaine

Capsaicin increases the lethality of cocaine

Cocaine's lethality increases during late gestation in the rat: A study of “critical periods” of exposure

OBJECTIVE: Cocaine-associated morbidities in pregnant women (e.g., abruptio placentae, hypertension, seizures) occur mostly during the final stages of gestation. The purpose of our study was to determine whether cocaine's toxicity and blood levels varied as a function of “critical periods” of exposure during gestation. STUDY DESIGN: To evaluate mortality rates, pregnant Long-Evans rats received subcutaneously 30, 40, or 50 mg/kg cocaine hydrochloride twice daily (C30, C40, and C50 groups) either during gestational days 7 to 13 (midgestation) or gestational days 14 to 20 (late gestation) ( n = 9 to 20 per group). Serum levels of the cocaine metabolite benzoylecgonine were examined in other groups of rats on either gestational day 13 (mid) or day 20 (late) in the C30 treatment condition ( n = 5 and 10 per group). RESULTS: There were no maternal mortalities in the midgestation groups at any dose. In contrast, the late-gestation groups showed a dramatic dose-dependent effect, with maternal mortality rates of 0%, 40%, and 72% in the C30, C40, and C50 groups. The late-gestation group had higher benzoylecgonine levels than the midgestation groups did. CONCLUSIONS: Late gestation was associated with higher maternal mortality rates and higher benzoylecgonine levels, indicating that some underlying physiologic change enhanced cocaine's toxicity as pregnancy progressed. This increased sensitivity to cocaine may be mediated by estrogen or progesterone, suggesting that the cocaine-abusing woman is at increased risk for cocaine-induced morbidities whenever levels of these hormones are elevated, such as during the final stages of pregnancy or possibly when taking oral contraceptives. (Am J Obstet Gynecol 1997;176:901-6.)

Role of dopamine D 1 receptors in cocaine lethality

One group of heterogeneously bred HS mice was assigned to test coadministration of the selective D 1 antagonist SCH 23390 with a dose of cocaine (95 mg/kg) that was observed to produce 80% lethality, whereas a second group was tested by cotreatment with the newly developed full-efficacy D 1 agonist dihydrexidine (DHX) and a dose of (60 mg/kg) cocaine previously shown to be nonlethal. The mice in the former group displayed decreased lethality going from 80% with coadministered vehicle to 15% after pretreatment with the highest dose (0.45 mg/kg) of SCH 23390. In the other group of mice there was no lethality seen when vehicle or 10 mg/kg DHX was coadministered with 60 mg/kg cocaine, but a dose-responsive increase in lethality with increasing DHX doses; the maximal lethality of 80% occurred when 25 mg/kg DHX was coadministered with cocaine. These results confirm the effects of D 1 antagonism decreasing cocaine lethality as reported previously when rats were used, and extend the findings to D 1 agonism; both observations evidence a role for the D 1 receptor in the lethal effects, be they central, cardiopulmonary, or anesthetic, of cocaine.

The effects of lidocaine pretreatment on cocaine neurotoxicity and lethality in mice.

To evaluate the effects of an antiarrhythmic dose of lidocaine on cocaine-induced ataxia, seizures, and death in mice. A randomized, controlled, blinded investigation was conducted using 220 female Swiss albino mice. Groups of 20 animals received intraperitoneal (IP) administration of either 31.6 mg/kg of lidocaine hydrochloride in 0.9% NaCl (ten animals) or an equal volume of 0.9% NaCl solution (ten animals). After 5 minutes, all the animals received IP cocaine in incremental doses ranging from 40 to 110 mg/kg. The animals were observed for ataxia, seizures, and death. The animals pretreated with lidocaine were compared with the control animals for the number of adverse effects from cocaine at each dose tested. Lidocaine significantly increased the overall incidences of cocaine-induced ataxia (p = 0.02) and seizures (p < 0.001). However, pretreatment with lidocaine offered protection against cocaine lethality (p < 0.0001). In this preexposure model, lidocaine protects mice against cocaine-induced lethality. These effects seem to be independent of convulsive activity. Further study is required to determine the safety and efficacy of lidocaine for the management of cocaine-induced arrhythmias.

Cocaine LD 50 in long-evans rats is not altered by pregnancy or progesterone

Progesterone increases cocaine's cardiovascular toxicity in sheep and rats. To determine whether progesterone enhances the lethality of cocaine, nonpregnant female rats were treated with either IM progesterone (P4) or vehicle, and pregnant rats (Preg) were untreated. The rats received one IP injection of cocaine at a dose between 25–75 mg/kg and were observed for seizures and/or death. All 62 rats that died did so within 17 min, preceded by seizures in 90.3%. Mean times-to-seizure and times-to-death, and mean lethal serum cocaine concentrations did not differ among groups. Serum progesterone levels (ng/ml ± SEM) at the time of death were different among groups: 24 ± 1.7 (C), 102 ± 6.4 (P4), and 139 ± 5.2 (Preg). Logistic regression dose/fatality curves, LD 50s, and LD 10s for the pregnant, progeste and control groups were not significantly different from one another. Though progesterone has enhanced cocaine's cardiac toxicity in some studies, it does not increase the risk of death from acute cocaine exposure in rats.

Urethane anesthesia reverses the protective effect of noncompetitive NMDA receptor antagonists against cocaine intoxication

The present experiments examined whether pretreatment with the noncompetitive N-methyl-D-aspartate (NMDA) receptor antagonists, MK-801 and dextrorphan, could antagonize cocaine-induced convulsions and lethality in conscious Sprague-Dawley (SD) rats and whether urethane anesthesia alters the observed interactions. Conscious, restrained male SD rats received continuous i.v. infusions of cocaine hydrochloride (1.25 mg/kg.min) until convulsions and death occurred. Cocaine doses of 21.2±1.8 and 29.5±2.5 mg/kg caused convulsions and death, respectively, in saline treated rats (n = 8). Convulsions were absent in MK-801 (1 mg/kg, i.v.;n = 8) pretreated rats; the lethal cocaine dose was 44.0±2.7 mg/kg (p<0.05). In contrast, urethane anesthesia (1.2 g/kg, i.p.) decreased the dose of cocaine required to cause toxicity, compared to that in saline controls (24.8 ± 0.8 mg/kg, n = 13), in MK-801 (2.0±0.3, n = 7; p<0.01) and in dextrorphan (25 mg/kg, i.v. ; 13.1±1.4, n = 6; p<0.01) pretreated rats. Pressor responses with little change in heart rate were evident during cocaine infusion in vehicle pretreated rats. Bradycardic responses were noted to cocaine in groups following NMDA receptor blockade. Reversal of the pressor response to cocaine was noted in MK-801 pretreated animals, while dextrorphan pretreatment moderated cocaine-induced increases in blood pressure. Ventilatory support protected against cocaine lethality in urethane anesthetized rats, indicating that respiratory failure is the proximate cause of death with cocaine infusion. However, artificially ventilated rats, pretreated with MK-801, were more sensitive (lethal cocaine dose, 76.6±8.0 mg/kg, n = 5) than vehicle pretreated rats (129.4±15.8 mg/kg, n = 6), indicating that MK-801 may increase both the respiratory and the cardiac toxicity of cocaine in urethane anesthetized rats. Interactions between NMDA receptors and cocaine are modified by urethane anesthesia.

Tetracaine protects against cocaine lethality in mice.

Tetracaine will enhance cocaine toxicity. Two hundred forty female Swiss albino mice weighing 27 to 45 g. Intraperitoneal injections of tetracaine and cocaine were given to groups of ten mice each in a controlled, blinded fashion. Either tetracaine or an equal volume of normal saline was given five minutes before one of six incremental doses of cocaine, ranging from 60 to 110 mg/kg. The experiment was repeated twice using two different doses of tetracaine: either an LD10 (40 mg/kg determined from preliminary studies), or one-twentieth of the dose of cocaine (which approximates the ratio used in tetracaine, epinephrine, and cocaine). Lethality was recorded at 24 hours. Lethality between groups was compared with a Wilcoxon sign-rank test. Tetracaine reduced cocaine lethality at all doses. This reduction in lethality was statistically significant at both tetracaine doses (P < .05). In the mouse, pretreatment with tetracaine significantly decreases cocaine lethality.

Effect of sex steroids on cocaine lethality in male and female mice

1. Endogenous sex steroid levels were altered in mice via gonadectomy, via physiological or supraphysiological doses of testosterone and/or estradiol, and via tamoxifen dosing to antagonize estrogens. 2. The role of sex hormones in susceptibility to cocaine lethality was examined via the response of mice after endocrine alterations to an intraperitoneal (i.p.) cocaine HCl (75 mg/kg). Incidence of deaths was significantly decreased only in sham-operated males receiving estradiol or tamoxifen and in ovariectomized or sham-operated females receiving doses of estradiol. 3. The levels of estradiol in both sexes appeared to be more influential than were levels of testosterone as a determinant of susceptibility to cocaine.

Modulation of the lethal effects of cocaine by cholinomimetics

In view of the toxicity of cocaine and recent reports on the antimuscarinic properties of cocaine, the present study evaluated the effects of manipulations in cholinergic neurotransmission on the lethal effects of cocaine. Physostigmine pretreatment significantly altered the lethality of cocaine, increasing the LD 50 from 82.5 mg/km to 96.9 mg/kg in male F344 rats. Atropine alone did not alter the lethal effects of cocaine at doses that are effective in preventing parasympathetic effects and lethality of oxotremorine. The prophylactic effect of physostigmine was not prevented by atropine. Neostigmine did not significantly affect the cocaine lethality dose-effect function. Both oxotremorine and (−)-nicotine were also devoid of protective actions. At higher doses, all of the cholinomimetics potentiated the lethal effects of cocaine. These results suggest that whereas cocaine lethality may be enhanced by stimulation of muscarinic receptors, low doses of physostigmine protect against lethality through actions at noncholinergic sites.

Antagonism of intravenous cocaine lethality in nonhuman primates.

(1980). Antagonism of Intravenous Cocaine Lethality in Nonhuman Primates. Clinical Toxicology: Vol. 16, No. 4, pp. 499-508.

Intravenous cocaine lethality in the rat

Summary A series of experiments was conducted in order to ascertain several factors related to the convulsant (lethal) effect of cocaine following intravenous administration to freely moving rats. The results of the initial study demonstrated that as the concentration of a cocaine infusion is increased, the time to death and the mean lethal dosage are significantly reduced. Pretreatment with non-depressing dosages of either alpha-methylparatyrosine, pimozide, or U-14,624 failed to alter either the convulsant action or the mean acute lethal dose of cocaine. These results suggest that the convulsant and/or lethal action of cocaine may not be mediated by catecholamines. Subacute intravenous treatment with cocaine for seven consecutive days resulted in a significant elevation in the mean lethal dose of cocaine. However, only half of the subjects survived this treatment regimen. In those subjects not surviving, evidence of sensitization to the convulsant action of cocaine was demonstrated.