Gamma-aminobutyric acid and alcohol actions: neurochemical studies of long sleep and short sleep mice

Effects of ethanol and pentobarbital on the GABA receptorchloride channel complex were evaluated in mice selected for differential sensitivity to the hypnotic effects of ethanol (long sleep and short sleep lines). 36Cl- influx, [35S]tbutylbicyclophosphorothionate (TBPS) and [3H]muscimol binding were measured in a membrane vesicle suspension (microsacs) from cerebellum or forebrain. Muscimol was found to be a more potent stimulator of 36Cl- flux in the LS cerebellum, as compared to the SS cerebellum, but a similar maximal level of uptake was achieved in the two lines. Muscimol displaced [35S]TBPS (a ligand for the convulsant site) from cerebellar microsacs, and LS mice were also more sensitive than SS mice to this action of muscimol. However, the number or affinity of high affinity [3H]muscimol binding sites did not differ between the lines. Physiologically relevant concentrations of ethanol (15-50 mM) potentiated muscimol stimulation of 36Cl- uptake in LS cerebellum but had no effect in SS cerebellum. Ethanol failed to alter stimulated chloride flux hippocampal microsacs from either line. Both the LS and SS lines responded similarly to pentobarbital potentiation of muscimol stimulated chloride uptake regardless of brain region. The demonstrated difference between the LS and SS mice in muscimol stimulated chloride uptake as well as in muscimol displacement of [35S]TBPS binding offers a biochemical explanation for the line differences in behavioral responses to GABAergic agents. Moreover, the findings suggest that genetic differences in ethanol hypnosis are related to differences in the sensitivity of GABA-operated chloride channels to ethanol.     

Relationship of membrane physical properties to alcohol dependence in mice selected for genetic differences in alcohol withdrawal

Genetic selection based on severity of withdrawal seizures following inhalation of ethanol vapor has produced two lines of mice, WSR (withdrawal seizure resistant) and WSP (withdrawal seizure prone), that differ markedly in withdrawal signs. In the present study, we report that these mice also differed in the severity of withdrawal seizures following consumption of an ethanol-containing liquid diet but did not differ in ethanol intake. In contrast to ethanol withdrawal seizures, the lines displayed similar sensitivity to electrical- or pentylenetetrazole-induced seizures. These results suggest that the lines differ in the development of physical dependence on ethanol rather than seizure sensitivity per se. Because decreased synaptic membrane fluidity has been associated with ethanol dependence, we used fluorescence polarization of diphenylhexatriene and trimethylammonium-diphenylhexatriene to evaluate membrane fluidity in WSP and WSR mice fed lab chow, an ethanol-containing liquid diet, or an isocaloric sucrose-containing liquid diet. Fluidity of brain synaptic membranes was identical for WSP and WSR mice fed lab chow. The control liquid diet did not alter membrane fluidity, and the ethanol diet decreased fluidity equally for WSP and WSR mice. Thus, the genetic difference in development of ethanol dependence found in these lines was not reflected in the physical properties of brain membranes.     

Specific ethanol withdrawal seizures in genetically selected mice

We are selectively breeding mice prone (WSP) and resistant (WSR) to ethanol withdrawal seizures assessed by handling induced convulsions (HIC). The possibility that differences between the lines in HIC scores are a result of differences in general CNS excitability not specific to ethanol withdrawal was examined. Using treatments which produce generalized seizures (electroconvulsive shock, strychnine, and flurothyl) and gamma amino-butyric acid (GABA) antagonists (picrotoxin, bicuculline, and pentylentetrazol), the ED50 for seizures was determined in the selected lines. In addition, the sensitivity of WSP and WSR mice to the anticonvulsant actions of ethanol against each treatment was determined. Neither the convulsant amperage 50 (CA50) for ECS nor the ED50 for any drug treatment differed for the selected lines. When ethanol (1.5 g/kg) was administered prior to ECS, there was a dramatic differential suppression of ECS in the lines: the CA50 of WSR mice was elevated 5-fold, whereas the CA50 of WSP mice increased only two fold. Ethanol pretreatment also elevated the ED50 for strychnine and flurothyl in WSR mice significantly more than WSP mice, but the line difference was smaller than for the anticonvulsant effect against ECS. The ED50s for the GABA antagonists were not different between the WSR and WSP lines after ethanol pretreatment. We conclude that genetic selection is producing lines of mice that differ specifically in the degree of seizure severity caused by withdrawal from ethanol physical dependence and not in generalized CNS excitability. An increased sensitivity to the anticonvulsant effects of ethanol against some convulsant treatments has appeared as a correlated response to selection in the WSR line.     

Correlation between the enhancement of flunitrazepam binding by GABA and seizure susceptibility in mice

Various populations of mice exhibit differential sensitivity to seizure-inducing agents. The relationship of seizure susceptibility to alterations in the GABA receptor complex was investigated in six different populations of mice consisting of four inbred strains (C57BL, DBA, C3H, and BALB) and two selected lines (long sleep and short sleep). Seizure activity was induced by intraperitoneal administration of the GAD inhibitor, 3-mercaptopropionic acid, and latencies to seizure onset and tonus were measured. In naive mice of the same populations, GABA enhancement of 3H-flunitrazepam binding was measured in extensively washed whole brain membranes at several GABA concentrations. Both differential seizure sensitivity to 3-mercaptopropionic acid and differential enhancement of 3H-flunitrazepam binding by GABA were observed in these six populations of mice. Correlational analyses indicated a positive correlation between the degree of GABA enhancement of 3H-flunitrazepam binding and resistance to the seizure-inducing properties of 3-mercaptopropionic acid. These data suggest that genetic differences in sensitivity to seizure-inducing agents that disrupt the GABAergic system may be related to differences in coupling between the various receptors associated with the GABA receptor complex.     

Ethanol and nitrous oxide produce withdrawal-induced convulsions by similar mechanisms in mice

Twenty generations of selective breeding were used to produce lines (strains) of mice which differ markedly from one another in ethanol physical dependence development as indexed by handling-induced convulsions (HIC) induced by withdrawal from ethanol. These withdrawal seizure prone (WSP) and withdrawal seizure resistant (WSR) selection lines now differ by over 10-fold in HIC scores after equivalent exposure to intoxicating levels of ethanol via inhalation. Since handling-induced convulsions can be readily elicited following withdrawal from nitrous oxide, we sought to determine if the very large differences in ethanol withdrawal-induced HIC bred into these selection lines would generalize to nitrous oxide. Following a 60 min exposure to 75% nitrous oxide (in O2), a greater than 10-fold difference in HIC scores, and a 2-fold difference in tremor incidence was seen upon withdrawal in WSP vs. WSR mice. These findings closely parallel those seen with ethanol, and demonstrate that a large degree of commonality exists in the genes and the mechanisms determining these withdrawal signs. HIC elicited by nitrous oxide withdrawal were readily suppressed by ethanol, and HIC elicited by ethanol withdrawal were promptly suppressed by 75% nitrous oxide in WSP mice. Nitrous oxide also suppressed HIC and tremor associated with nitrous oxide withdrawal.     

Ethanol and diazepam withdrawal convulsions are extensively codetermined in WSP and WSR mice

Selective breeding was used to produce lines of mice which differ markedly in their genetically-mediated vulnerability to handling-induced convulsions (HIC) associated with the ethanol withdrawal syndrome. These are known as the ethanol withdrawal seizure prone (WSP) and withdrawal seizure resistant (WSR) selection lines. As a result of 5 generations of selective breeding with ethanol, a 3.4-fold difference between WSP and WSR mice was seen in HIC associated with ethanol withdrawal. When diazepam was used as the dependence-producing drug, a 2.4-fold difference emerged. After 6 more generations of selective breeding with ethanol, an approximate 10-fold difference was seen with ethanol, while with diazepam, this difference in HIC scores was also about 10-fold. This close parallel between ethanol and diazepam indicates that physical dependence on both drugs, as indexed by handling-induced convulsions, is extensively codetermined by the same genes, and thus by the same mechanisms, in these selectively-bred mice.     

Potential pleiotropic effects of Mpdz on vulnerability to seizures

We previously mapped quantitative trait loci (QTL) responsible for approximately 26% of the genetic variance in acute alcohol and barbiturate (i.e., pentobarbital) withdrawal convulsion liability to a < 1 cM (1.8 Mb) interval of mouse chromosome 4. To date, Mpdz , which encodes the multiple PSD95/DLG/ZO-1 (PDZ) domain protein (MPDZ), is the only gene within the interval shown to have allelic variants that differ in coding sequence and/or expression, making it a strong candidate gene for the QTL. Previous work indicates that Mpdz haplotypes in standard mouse strains encode distinct protein variants (MPDZ1-3), and that MPDZ status is genetically correlated with severity of withdrawal from alcohol and pentobarbital. Here, we report that MPDZ status cosegregates with withdrawal convulsion severity in lines of mice selectively bred for phenotypic differences in severity of acute withdrawal from alcohol [i.e., High Alcohol Withdrawal (HAW) and Low Alcohol Withdrawal (LAW) lines] or pentobarbital [High Pentobarbital Withdrawal (HPW) and Low Pentobarbital Withdrawal (LPW) lines]. These analyses confirm that MPDZ status is associated with severity of alcohol and pentobarbital withdrawal convulsions. Using a panel of standard inbred strains of mice, we assessed the association between MPDZ status with seizures induced by nine chemiconvulsants. Our results show that MPDZ status is genetically correlated with seizure sensitivity to pentylenetetrazol, kainate and other chemiconvulsants. Our results provide evidence that Mpdz may have pleiotropic effects on multiple seizure phenotypes, including seizures associated with withdrawal from two classes of central nervous system (CNS) depressants and sensitivity to specific chemiconvulsants that affect glutaminergic and GABAergic neurotransmission.     

Effect of chronic ivermectin treatment on GABA receptor function in ethanol withdrawal-seizure prone and resistant mice

Ivermectin, a potent, effective anthelmintic, is easy to administer, has a broad spectrum of action and a wide safety margin. However, no testing has been done in hosts genetically selected for seizure susceptibility which may be more sensitive to the effects of ivermectin than other animals. This was done in the present experiments with seizure prone and seizure resistant mice infested with Syphacia obvelata (pinworm). These subjects were treated daily with oral ivermectin in their drinking water every other week for six weeks, for a total of 21 days. The treatment cleared the mice of the pinworm infestation, but did not alter the seizure susceptibility or binding parameters of [3H]flunitrazepam in either of the selected lines.     

Sleep-time variation for ethanol and the hypnotic drugs tribromoethanol, urethane, pentobarbital, and propofol within outbred ICR mice.

To evaluate the phenotypic variation within a commercial outbred mouse stock, we examined sleep-time (or duration of loss of righting reflex) of outbred ICR mice after i.p. injection of ethanol (4.0 g/kg of body weight), urethane (1.3 g), tribromoethanol (250 mg), and pentobarbital (60 mg), and after i.v. injection of propofol (30 mg). We observed high-grade individual differences in sleep-time that ranged from 0 to 179 min, 83.1 +/- 4.3 (mean and SEM of 100 mice) for ethanol; 0 to 169 min, 64.5 +/- 3.1 for pentobarbital; 0 to 160 min, 36.6 +/- 3.6 for urethane; 0 to 120 min, 21.5 +/- 2.2 for tribromoethanol; and 3 to 20.5 min, 7.1 +/- 0.3 for propofol. This extensive phenotypic variance within the outbred stock was as great as the variation reported among inbred strains or selected lines, and the varied susceptibility within the colony was inherited by Jcl:ICR-derived inbred strains IAI, ICT, IPI, and IQI. The range of sleep-time variance for ethanol, pentobarbital, urethane, tribromoethanol, and propofol within four-way cross hybrid Jcl:MCH(ICR) mice was 86.6%, 63.3%, 124%, 61.0%, and 53.1% that of outbred Jcl:ICR mice, respectively. The present study indicates that phenotypic variance within an outbred Jcl:ICR stock was at high risk for susceptibility to the drugs that depress the central nervous system and that Jcl:ICR-derived inbreds may be an excellent source of animal models for studying the anesthesia gene.     

Marked variation in diazepam sensitivity in Swiss albino mice

Using the righting reflex as the critical level, sleep was measured in Swiss albino mice at a dose of 35 mg/kg diazepam, i.p. Sleep times varied markedly from zero to 120 min with a mean +/- s.d. of 44 +/- 37 (N = 202). The distribution is skewed to the left with a coefficient of skewness of 0.33 +/- 0.17. The sleep times of the two sexes, when analyzed separately, showed similar range, mean and s.d., except that the distribution tended to be more clearly bimodal in males than in females. These animals also exhibited marked variations in their response to either ethanol (4 g/kg) or pentobarbital (45 mg/kg). The diazepam sleep time failed to correlate with the ethanol sleep time. Significant correlation, however, was obtained between diazepam and pentobarbital sleep times. On further analysis with least-squares fit to a straight line, the data yielded a line with a slope of 0.16; thus despite the correlation reaching a significant level, there is no significant difference in the pentobarbital sleep times between mice that have the longest or the shortest diazepam sleep times. By monitoring the plasma and brain levels of diazepam and N-desmethyldiazepam in mice at awakening, it was found that the variations in sleep time cannot be explained by individual differences in drug disposition. The phenomenon is discussed in terms of individual variations in diazepam sensitivity and the possibility of development of tolerance to diazepam almost immediately after diazepam administration.     

High genetic susceptibility to ethanol withdrawal predicts low ethanol consumption

C57BL/6J (B6) inbred mice are well known to drink large amounts of alcohol (ethanol) voluntarily and to have only modest ethanol-induced withdrawal under fixed dose conditions. In contrast, DBA/2J (D2) mice are ``teetotallers' and exhibit severe ethanol withdrawal. Speculation that an inverse genetic relationship existed between these two traits was substantiated by meta-analysis of existing data collected in multiple genetic models, including large panels of standard and recombinant inbred strains, their crosses, and selectively bred mouse lines. Despite methodological differences among laboratories in measurement of both preference drinking and withdrawal, a nearly universal finding was that genotypes consuming large amounts of 10% ethanol (calculated as g/kg/day) during two-bottle choice preference drinking were genetically predisposed to low withdrawal scores in independent studies after either acute or chronic ethanol treatment. Conversely, low-drinking genotypes had higher withdrawal severity scores. The genetic relationship appears to be strongest in populations derived from B6 and D2, where data from more genotypes (BXD RIs, B6D2F₂s, BXD RI F₁s, and B6D2F₂-derived selectively bred lines) were available for analysis. Gene mapping studies in these populations identified four chromosome regions [on Chromosomes (Chrs) 1, 2, 4, and 15] where genes might potentially influence both traits. Among genotypes with greater genetic diversity (for example, a panel of standard inbred strains or selectively bred lines), the relationship was less pronounced. Thus, reduced susceptibility to the development of high alcohol use may be supported by increased genetic susceptibility to ethanol withdrawal symptoms. Received: 15 September 1998 / Accepted: 8 October 1998     

Effects of lorazepam tolerance and withdrawal on GABAA receptor operated chloride channels in mice selected for differences in ethanol withdrawal severity.

Withdrawal seizure prone (WSP) and withdrawal seizure resistant (WSR) mice were treated with 5 mg/kg lorazepam for 7 days via implanted osmotic mini pumps. Following chronic drug treatment, brains were assayed for GABA-mediated chloride flux (GABA-Cl-). Under control (drug naive) conditions, brain membranes prepared from WSP and WSR lines did not differ in flunitrazepam or ethanol stimulation of GABA-mediated 36Cl- uptake, but the WSP lines were more sensitive to inhibition of 36Cl- flux by the inverse agonist, FG-7142. Membranes from lorazepam tolerant WSP and WSR mice were resistant to flunitrazepam- and ethanol-stimulation of GABA-Cl-. Withdrawal from chronic treatment, by an acute injection with the benzodiazepine antagonist RO15-1788, returned flunitrazepam stimulation of GABA-Cl- to near control levels in WSR membranes but not in WSP membranes and restored ethanol modulation of the channel to control levels in both lines. Inhibition of chloride flux by the benzodiazepine partial inverse agonist, FG-7142 was greater in membranes from WSP mice compared with WSR mice. Tolerance to lorazepam increased sensitivity of the WSR membranes to FG-7142 without altering the response in the WSP line. Again, withdrawal restored the Cl- flux response to FG-7142 back to near control levels. Lorazepam tolerance lowered [3H]-flunitrazepam binding affinity slightly only in the WSR strain with no change in binding number. Withdrawal from chronic lorazepam treatment produced no significant change in binding affinity or number. The initial genotypic differences in benzodiazepine inverse agonist sensitivity, may be related to the selection for withdrawal seizure severity. Chronic administration of lorazepam reduces the coupling between the benzodiazepine agonist site and the chloride channel and concomitantly increases coupling between the channel and the inverse agonist site, while withdrawal resets the receptor coupling back to control response levels. However, for the WSP line, this drug environment dependent shift in channel coupling bias appears to be deficient compared with the WSR line.     

Targets for ethanol action and antagonism in loop 2 of the extracellular domain of glycine receptors

The present studies used increased atmospheric pressure in place of a traditional pharmacological antagonist to probe the molecular sites and mechanisms of ethanol action in glycine receptors (GlyRs). Based on previous studies, we tested the hypothesis that physical–chemical properties at position 52 in extracellular domain Loop 2 of α1GlyRs, or the homologous α2GlyR position 59, determine sensitivity to ethanol and pressure antagonism of ethanol. Pressure antagonized ethanol in α1GlyRs that contain a non-polar residue at position 52, but did not antagonize ethanol in receptors with a polar residue at this position. Ethanol sensitivity in receptors with polar substitutions at position 52 was significantly lower than GlyRs with non-polar residues at this position. The α2T59A mutation switched sensitivity to ethanol and pressure antagonism in the WTα2GlyR, thereby making it α1-like. Collectively, these findings indicate that (i) polarity at position 52 plays a key role in determining sensitivity to ethanol and pressure antagonism of ethanol; (ii) the extracellular domain in α1- and α2GlyRs is a target for ethanol action and antagonism and (iii) there is structural-functional homology across subunits in Loop 2 of GlyRs with respect to their roles in determining sensitivity to ethanol and pressure antagonism of ethanol. These findings should help in the development of pharmacological agents that antagonize ethanol.     

Factors affecting actions of ethanol on GABA-activated chloride channels.

Effects of ethanol in vitro on membrane vesicles (microsacs prepared from mouse cerebral cortex) were evaluated by monitoring 36Cl- influx. Different assay parameters were tested to determine increased or decreased action of ethanol on GABA-activated chloride channels. The ability of 30 mM ethanol to augment 36Cl- flux was seen at 0 degrees C, in the absence of GABA ("direct" action of ethanol), and at 34 degrees C in the presence of GABA, using two different assay procedures. Picrotoxin blocked the direct effects of ethanol (at 0 degrees C) suggesting GABAa involvement. Endogenous GABA in the medium surrounding the microsacs was assayed at different temperatures both in the presence and absence of GABA and ethanol. The direct effect of ethanol did not appear to involve the action of endogenous GABA. In addition to temperature effects on the assay, time of membrane storage also influenced ethanol action. Microsacs stored on ice for 2 hours or more lost their ability to respond to ethanol but not to GABA, pentobarbital or flunitrazepam. When these drugs were tested on membranes from mice that had been sacrificed by cervical dislocation as opposed to decapitation, ethanol did not augment GABA-stimulated chloride flux. The method of sacrifice did not influence the response to GABA, pentobarbital or flunitrazepam.     

Micro-spatial population differentiation in activity of glycerol-3-phosphate oxidase (GPO) from mitochondria of Drosophila melanogaster

Replicate mass-bred laboratory populations of D. melanogaster were derived from females collected in the Tahbilk winery cellar and from females collected outside but from within two kilometres of the cellar. When mitochondrial extracts from larvae were assayed for specific activity of glycerol-3-phosphate oxidase the cellar populations had levels only 50% of those from the outside area, confirming an earlier report of such a difference among isofemale lines derived from these same areas. This micro-spatial differentiation occurred when larvae were raised on a medium supplemented with both sucrose (5% w/v) and ethanol (4% v/v), known to effect high GPO activity, but was not detected when the larvae were raised on unsupplemented medium.A heritable basis for larval GPO activity variation was confirmed in a set of 32 isogenic second chromosome substitution lines and measured in a subset of 4 of these lines about 25 generations later. A reciprocal cross using two isogenic substitution lines with the highest and lowest activities suggested the difference was attributable to genes acting additively and that there were no maternal or paternal effects. The detection of a collection site difference in GPO enzyme activity in the isogenic lines suggests that polymorphic variation on the second chromosome is responsible for the differentiation at the winery.Variation in adult GPO activity did not show a dependence on the winery location from where the isogenic lines were derived nor was there an effect of line. Adult GPO activity was significantly higher than that detected in larval tissues and did not show a dependence on the sugar/ethanol level in the growth medium.     

Modulation of γ-Aminobutyric AcidA Receptor-Operated Chloride Channels by Benzodiazepine Inverse Agonists Is Related to Genetic Differences in Ethanol Withdrawal Seizure Severity

To determine whether genetic differences in development of ethanol dependence are related to changes in gamma-aminobutyric acidA (GABAA) receptor function, we measured 36Cl- uptake by brain cortical membrane vesicles from withdrawal seizure prone and withdrawal seizure resistant (WSP/WSR) mice treated chronically with ethanol. Muscimol-stimulated chloride flux was not different between WSP and WSR mice before or after ethanol treatment. Also, augmentation of muscimol action by flunitrazepam or inhibition of muscimol action by the inverse agonists Ro 15-4513 (ethyl-8-azido-5,6-dihydro-5-methyl-6-oxo-4H-imidazo[1,5a]- [1,4]benzodiazepine-3-carboxylate) and methyl-6,7-dimethoxy-4-ethyl-beta-carboline-3-carboxylate (DMCM) was not different for ethanol-naive WSP and WSR mice. However, chronic ethanol administration enhanced the inhibitory actions of DMCM and Ro 15-4513 on membranes from WSP but not WSR mice. Conversely, chronic ethanol treatment attenuated the action of flunitrazepam on membranes from WSR but not WSP mice, suggesting that the actions of benzodiazepine agonists and inverse agonists are under separate genetic control. These genetic differences in actions of DMCM and Ro 15-4513 indicate that sensitization to benzodiazepine inverse agonists produced by chronic ethanol treatment may be related to development of withdrawal seizures and suggest that differences in the GABA/benzodiazepine receptor complex represent alleles that have segregated during the selection of the WSP/WSR mice.     

Selection for ethanol tolerance in two populations of Drosophila melanogaster segregating alcohol dehydrogenase allozymes.

Selection for ethanol tolerance was equally successful in two populations of D. melanogaster in both of which the frequency of AdhF was 0.5 at the start of the experiment. Increased tolerance to ethanol was not invariably associated with increased frequencies of AdhF. In one population alcohol dehydrogenase (ADH) activity was significantly higher in three of the four selected sublines compared with their controls but there was no difference in activity between the selected and control sublines in the second population. The level of ADH activity in the control and selected lines was significantly correlated with the frequency of AdhF, but not with ethanol tolerance. These results show that adaptation to environmental alcohols in populations of D. melanogaster can be independent of the ADH system.     

Preference for ethanol in feeding and oviposition in temperate and tropical populations of Drosophila melanogaster

The natural habitat of Drosophila melanogaster Meigen (Diptera: Drosophilidae) is fermenting fruits, which can be rich in ethanol. For unknown reasons, temperate populations of this cosmopolitan species have higher ethanol resistance than tropical populations. To determine whether this difference is accompanied by a parallel difference in preference for ethanol, we compared two European and two tropical African populations in feeding and oviposition preference for ethanol‐supplemented medium. Although females of all populations laid significantly more eggs on medium with ethanol than on control medium, preference of European females for ethanol increased as ethanol concentration increased from 2 to 6%, whereas that of African females decreased. In feeding tests, African females preferred control medium over medium with 4% ethanol, whereas European females showed no preference. Males of all populations strongly preferred control medium. The combination of preference for ethanol in oviposition, and avoidance or neutrality in feeding, gives evidence that adults choose breeding sites with ethanol for the benefit of larvae, rather than for their own benefit. The stronger oviposition preference for ethanol of temperate than tropical females suggests that this benefit may be more important in temperate populations. Two possible benefits of ethanol for which there is some experimental evidence are cryoprotection and protection against natural enemies.     

Drosophila-Host Genetic Control of Susceptibility to Drosophila C Virus

Interactions between Drosophila C virus (DCV) and its natural host, Drosophila melanogaster, were investigated using 15 geographical population samples infected by intraabdominal inoculation. These strains derived from natural populations of D. melanogaster differed in susceptibility to the DCV(C). One strain was ``partially tolerant'. Isofemale lines obtained from one susceptible and one partially tolerant strain were studied. The partially tolerant phenotype was dominant, and there was no difference between F(1) progeny of direct and reciprocal crosses. Analysis of F(2) progeny showed that neither sex-linked genes nor maternal effects are involved in susceptibility to DCV(C). The partially tolerant strain phenotype was dominant and segregated with chromosome III. Two nonexclusive hypotheses are proposed to explain chromosome III gene action.     

Effect of body temperature on acute ethanol toxicity

We studied the effect of elevated rectal temperature on ethanol toxicity and the effect of ethanol on the mean lethal temperature (LT50). The rectal temperature was maintained at a preselected level for 4 h. Ethanol (23% w/v) was injected i.p. In control mice anesthetized with pentobarbital, the 4 h LT50 was 41.8 ± 0.1°C (mean ± standard error). In mice which had received a non-lethal ethanol dose (6 g/kg), the LT50 was decreased to 39.0 ± 0.2°C (p< 0.001). In control mice, whose temperature dropped, the 4 h ethanol LD50 was 8.5 ± 0.8 g/kg. If the rectal temperature of the mice was maintained instead at a maximum of 38.5 – 40°C, the LD50 was decreased to 5.7 ± 0.8 g/kg (p < 0.02). The results show that ethanol increases the susceptibility of mice to hyperthermic damage, and conversely, that hyperthermia increases the toxicity of ethanol.