Peter K. Gessner

Isobolographic analysis of interactions: an update on applications and utility

Isobolographic analysis provides a fundamental basis for assessing whether biological responses induced by mixtures of agents are greater, equal or smaller than would have been expected on the basis of the individual activities of the component agents and the concept of dose additivity. Limited in its direct application to binary mixtures, isobolographic analysis provides a conceptual framework and an unambiguous terminology, as well as an algebraic paradigm for the analysis of the interaction of ternary and higher order mixtures. A library of examples generously illustrated graphically is provided to facilitate the understanding of the methodology and serve as a guide for investigators who are unfamiliar with the approach. Also discussed are the theoretical derivation of the isobologram, the representation of various dosage combinations, the derivation of the principle of dose additivity, supra-additivity, infra-additivity, antagonism, the methods for probit analysis of mixture potency, effect addition and the consequences of peak effect coincidence in time or lack thereof, and the role of isobolographic analysis in the various aspects of dose-response surface methodology.

Dose-dependent consequences of cocaine on pregnancy outcome in the Long-Evans rat

The number of obstetric patients abusing cocaine has increased dramatically in recent years. To better understand its effect on pregnancy and to establish the LD50s for maternal and fetal fatalities, the dose-dependent effects of cocaine on pregnancy outcome were investigated in the Long-Evans rat. Pregnant animals were given either saline or 40, 50, 60, 70, 80, or 90 mg/kg cocaine hydrochloride from gestation days 7 to 19 inclusive. An additional group was non-treated and had ad lib access to food and water. Animals were sacrificed on gestation day 20 and the fetuses were examined. Despite treatment during the major periods of organogenesis and brain development, few congenital abnormalities were observed. There were, however, dose-dependent effects on maternal weight gain, maternal food and water consumption, fetal weight, maternal and fetal fatalities, fetal edema, abruptio placentae and cephalic hemorrhages. Despite suppression of maternal weight gain, there was preservation of fetal weights at cocaine doses up to and including 80 mg/kg/day, suggesting some protection of fetal growth. In addition to providing information on the gestational effects of cocaine in the rat, the present study provides information useful in guiding the selection of cocaine doses for subsequent behavioral teratology studies.

The interactive effects of alcohol and cocaine on maternal and fetal toxicity in the Long-Evans rat

The number of obstetric patients with polydrug abuse problems has increased substantially in recent years. One of the most common drug combinations is alcohol and cocaine. The effect of this drug combination on pregnancy is, therefore, of interest. Consequently, the present study investigated the relative and interactive effects of these two drugs on pregnancy outcome in an animal model. Alcohol and cocaine were administered, both separately and in combination, to separate groups of pregnant Long-Evans rats from gestation day 7-19. Animals were then sacrificed and examined on gestation day 20. Control animals were given vehicle only or were nontreated. The isobolographic method was used to evaluate the effects of the alcohol-by-cocaine interaction on select maternal and fetal variables. This method of analysis indicated that alcohol and cocaine had interactive effects that were linearly additive for some variables and infraadditive for others. In general, the results suggest that the alcohol-plus-cocaine drug combination poses a greater risk to pregnancy than either drug alone.

The disulfiram-ethanol reaction

The disulfiram-ethanol reaction (DER) is the name given to a syndrome experienced by individuals who, following treatment with disulfiram (DSSD), ingest ethanol. The DER is decidedly unpleasant. Accordingly, ever since the discovery of this syndrome, efforts have been made to utilize DSSD in aversive therapies of alcoholism, though the early therapeutic strategies employed differed markedly from the current ones (section 10.2). Partly as a result of this, a great deal has been learned regarding the pharmacological phenomena and toxicities characteristic of the human DER (section 10.3).

Failure of diphenylhydantoin to prevent alcohol withdrawal convulsions in mice.

Abstract Alcohol withdrawal reactions were induced in mice using an established technique. Mice, administered 1.0 mmole/kg pyrazole daily were exposed for 3 days to 10 mg/l ethanol vapor and withdrawal was brought about by discontinuation of the exposure. Diphenylhydantoin administered in doses of 12, 20 or 50 mg/kg by either of two routes (i.p. or p.o.), failed to have any discernable effect on the withdrawal. A 100 mg/kg dose of diphenylhydantoin increased seizure scores. Chloral hydrate administration in doses of 175, 244 or 350 mg/kg, on the other hand, lowered seizure scores in a manner which was dose related and prompt. These findings suggest that the clinical use of diphenylhydantoin for control of the seizures seen in alcohol withdrawal should be re-evaluated. The seizure scores of mice not exposed to ethanol but administered 1.0 mmole/kg pyrazole daily for 3 days were negligible and well within values obtained with naive control mice not exposed to any pharmacological agent.

Pharmacokinetics of paraldehyde disposition in the neonate

The pharmacokinetic parameters controlling paraldehyde elimination were determined in nine infants infused with paraldehyde at the rate of 150 mg/kg/hr in a 5% solution in 5% dextrose for the treatment of status epilepticus. The mean +/- SEM values for the observed parameters were as follows: rate constant for the disposition of paraldehyde 0.0680 +/- 0.0071 hr,-1 half-life 10.2 +/- 1.0 hr; volume of distribution 1.73 +/- 0.20 L/kg; clearance 0.121 +/- 0.023 L/hr/kg. Phenobarbital administration prior to or within 24 hours of the cessation of paraldehyde infusion decreased both paraldehyde clearance and volume of distribution in a manner linearly related to the logarithm of the phenobarbital dose. The rate constant for paraldehyde elimination was decreased as a linear function of the logarithm of the combined dose of administered phenobarbital and phenytoin. No acetaldehyde was detected in any blood samples. Paraldehyde administration was not correlated with any adverse reactions or toxicities.

Antagonism of the tranylcypromine--meperidine interaction by chlorpromazine in mice.

Abstract P.K. GESSNER. Antagonism of the tranylcypromine-meperidine interaction by chlorpromazine in mice, European J. Pharmacol 22 (1973) 187–190. Chlorpromazine protected mice against the toxicity resultant from administration of tranylcypromine and mepertidine 4 hr apart in a 3 10 ratio by weight. Doses of 3 mg/kg Chlorpromazine administered i.p. 1 hr prior to meperdine completely reserved the mortality seen after the i.p. administration of 21 mg/kg tranylcypromine and 70 mg/kg meperidine. Chlorpromazine was also found to protect mice significantly against meperidine toxicity. Administration of chlorpromazine, 10 mg/kg, 3 hr after tranylcypromine resulted in a marked fall in body temperature and a failure of meperidine. administered 1 hr later, to bring about the hyperthermic response seen in controls.

Metabolism of paraldehyde to acetaldehyde in liver microsomes. Evidence for the involvement of cytochrome P-450.

A concentration-dependent acetaldehyde (AcH) generation was observed when paraldehyde was incubated with the mouse liver microsomal fraction. The process, which exhibited a requirement for oxygen and NADPH and was inhibited by carbon monoxide, was found to have a Km of 17.9 mM with respect to paraldehyde and a Vmax of 40.1 nmoles/mg protein/min with respect to AcH formation. NADH was much less effective as an electron donor than NADPH, though a more than additive increase in AcH generation was observed when both of these nucleotides were added to the incubation. The rate of microsomal AcH generation from paraldehyde was increased 2.5-fold by pretreatment of the mice with phenobarbital but only 0.6-fold by pretreatment with 3-methylcholanthrene. Pretreatment with 2-diethylaminoethyl-2,2-diphenylvalerate hydrochloride (SKF-525A) resulted in 54% inhibition of the reaction rate. Addition of metopirone to the incubation inhibited AcH generation in a concentration-related fashion, the inhibition being greatest, proportionately, in microsomes from phenobarbital-pretreated animals. The above results conclusively indicate the involvement of cytochrome P-540 mixed function oxidase in the formation of AcH from paraldehyde by mouse liver microsomes. It is also postulated that this process may be accomplished in the reaction analogous to O-dealkylation.

Metabolism of disulfiram and diethyldithiocarbamate

The first and rapid step in the metabolism of disulfiram (DSSD) is the reduction of its disulfide bond. This biotransformation can be effected by endogenous thiols, sulfhydryl groups of proteins (section 5.2), or reduced forms of redox cycling metal ions (section 5.4). Moreover, such reactions can occur in blood, liver and other tissues. The eventual end result of these reactions is the conversion of both the DS moieties of DSSD to diethyldithiocarbamic acid (DSH). The sulfhydryl groups of thiols and proteins react with DSSD to yield immediately one equivalent of DSH. The second equivalent of DSH becomes available when the DS residue-containing mixed disulfide is subsequently reduced. In the case of proteins the latter reduction can be brought about by a vicinal sulfhydryl group, if one is present. This results in the formation of an intramolecular disulfide bond between the protein’s vicinal sulfhydryls while DSH is released (section 8.1). Reducing metal ions react with DSSD to form chelates from which DSH can be later displaced (section 5.4).

Effects of meperidine on operant behavioral thermoregulation in mice under conditions of symmetrical thermal drive

The effects of meperidine on operant behavioral thermoregulation were investigated using a convective thermal controller and mice trained to alternate at will the thermal-drive condition, changing it from an air flow of 15° C to one of 45° C, and vice versa. Administration of 15 mg/kg meperidine resulted in significantly lower response rates, a significantly larger fraction of time spent in one (mostly cold) drive condition, and significantly lower body temperatures than administration of saline. In a second experiment the animal was automatically returned to the hot-drive condition every 3 min unless it had been exposed to this condition in the previous 0.4 min although otherwise it remained free to alternative drive conditions at will. Under these conditions, meperidine-treated animals also spent significantly more time in cold drive and had significantly lower body temperatures than control animals, in spite of lower response rates. Pretreatment with 4.5 mg/kg tranylcypromine (4 h prior) did not significantly alter the effects of meperidine administration in either experiment.