Short-sleep Mice Research Articles

The effects of chronic ethanol ingestion on ethanol binding to hepatic cytochrome P-450 and on certain hepatic and renal parameters in the "long sleep" and "short sleep" mouse.

Male mice selected for genetic differences in ethanol-induced sleep time, thereby designated long sleep (LS) an short sleep (SS), were treated with the Lieber-DeCarli liquid diet for 25 days. This chronic ethanol treatment produced an increase in liver/body weight and kidney/body weight in SS mice only. In addition, chronic ethanol treatment produced significant increases in both LS and SS treated mice in in vivo ethanol elimination, hepatic cytochromes P-450 and B5, NADPH cytochrome c reductase and hepatic and renal 7-ethoxycoumarin O-de-ethylase activity. Genotypic differences were observed in the magnitude of response of microsomal ethanol oxidation per mg of microsomal protein (SS greater than LS). Further, control LS and SS mice possessed substantially different activity of renal 7-ethoxycoumarin O-de-ethylase. Both lines exhibited similar induced renal 7-ethoxycoumarin O-de-ethylase activity after chronic ethanol ingestion. Ethanol binding spectra produced when ethanol was added to hepatic microsomes were examined using double reciprocal plots. Chronic ethanol ingestion produced genotypically related (LS greater than SS) increases in the absorbance change maximum per mg of microsomal protein. No significant changes in the spectral dissociation constant or absorbance change maximum per nM cytochrome P-450 were observed following ethanol treatment.

Ethanol-induced depressions in cerebellar and hippocampal neurons of mice selectively bred for differences in ethanol sensitivity: An electrophysiological study

The recently discovered profound differential sensitivity of cerebellar Purkinje (P) cells in long-sleep (LS) versus short-sleep (SS) mice to the depressant effects of locally applied ethanol was extended in this study. First, the sensitivity of Purkinje neurons from HS mice (an outbred stock of mice from which the LS and SS lines were derived), was found to be almost exactly intermediate between the values for the long-sleep and short-sleep animals. Second, no differential sensitivity in long-sleep versus short-sleep hippocampal pyramidal neurons was observed. This was true using both spontaneous and evoked activity. Third, no differential sensitivity of P cells was seen in long- versus short-sleep mice with local application of halothane. Taken together with previous reports, these data strongly suggest that whatever genetically determined central nervous alterations result in the differential soporific effects of ethanol in the two (LS and SS) mouse lines, such alterations are brain region- and depressant drug-specific rather than generalized.

ANAESTHETIC REQUIREMENT IN MICE SELECTIVELY BRED FOR DIFFERENCES IN ETHANOL SENSITIVITY

Anaesthetic requirements for nitrous oxide, enflurane and isoflurane were determined in mice selectively bred for their susceptibility ("long-sleep" mice) or resistance ("short-sleep" mice) to alcohol. Nitrous oxide and enflurane requirements, measured by the rolling-response test, were 34 and 20% greater, respectively, in short-sleep mice than in long-sleep mice. Although isoflurane requirement was 39% greater when measured by the tail-clamp test, it was not significantly different when measured by the rolling-response test. The greater anaesthetic requirement for short-sleep mice was not associated with a different synaptic membrane phospholipid, fatty acid or cholesterol composition.

Withdrawal-like signs induced by a single administration of ethanol in mice that differ in ethanol sensitivity.

These experiments studied changes produced by a hypnotic dose of ethanol in the LS and SS lines of mice, which differ in ethanol sensitivity. In the first experiment, animals were injected either with ethanol or saline, and activity and seizure susceptibility measured 7-9 h later when blood levels of ethanol would have reached zero. Ethanol-treated mice of both genetic lines were less active in an open field test and more susceptible to clonic convulsions induced by flurothyl than saline-injected controls. There was no difference in the magnitude of these changes in the two lines. In the control condition SS (short-sleep) mice were more active than LS (long-sleep) mice, and more susceptible than LS mice to myoclonic but not to clonic seizures. The effect of the ethanol injection on body temperature was evaluated in separate groups of animals. LS mice showed a more pronounced hypothermia than SS mice when temperature was measured 2 h after injection. Six hours after injection, SS mice exhibited a small but statistically significant overshoot in temperature, after which they again became hypothermic with respect to controls; hyperthermia was not observed in LS animals.

Brain neurotransmitter receptor systems in mice genetically selected for differences in sensitivity to ethanol

The density of receptors for neurotransmitters and receptor-mediated cellular responsiveness were determined in tissues from two lines of mice selectively bred for differences in sleep time after ethanol administration. The studies were undertaken to ascertain in differences in neurotransmitter receptors between the lines might explain any of the differences seen in initial sensitivity to ethanol. The results were compared with those obtained in studies using the parent heterogeneous stock (HS). The animals used in the current experiments did not receive ethanol. β-Adrenergic receptor density, as measured by the binding of ( 125I)-iodohydroxybenzylpindolol, was lower in cortices of alcohol-sensitive long-sleep mice (LS) as compared to the short-sleep (SS) animals or to HS controls. The densities of β 1 and β 2-adrenergic receptors were lower by approximately the same amount. The results were tissue specific in that there was no difference in the density of β-adrenergic receptors in the cerebellum. Cyclic AMP accumulation in response to isoproterenol was not significantly different in the cortex of LS mice despite the decrease in β-adrenergic receptors. No differences were found in 3H-spiroperidol binding to dopamine receptors in the striatum or in 3H-quinuclidinylbenzilate binding to muscarinic cholinergic receptors in the cortex, striatum or hippocampus. Dopamine-stimulated adenylate cyclase activity was, how- ever, lower in striata of SS mice. The affinities of the receptors for the various ligands studied were the same in the three lines of mice.

Genetically mediated responses of microsomal ethanol oxidation in mice.

Long-sleep (LS) and Short-sleep (SS) male mice were treated chronically with alcohol for 30 days. Ethanol treated mice were withdrawn from alcohol diet at 8, 12, 16, 20, 24 and 30 days of treatment for the assessment of metabolic tolerance and numerous parameters associated with the microsomal cytochrome P-450 complex. Up to 20 days of ethanol treatment, LS and SS mice displayed nearly the same enhancement of in vivo ethanol elimination rates. AT 24 and 30 days of ethanol treatment, LS mice were found to have significantly greater ethanol elimination rates than SS mice chronically treated with alcohol. The induction of microsomal cytochrome P-450 content and microsomal ethanol oxidation paralleled the acquisition of metabolic tolerance with LS mice displaying significantly greater values of both microsomal parameters at 24 and 30 days of treatment. Aniline hydroxylase was maximally induced (2.5 fold) in both LS and SS mice by 16 days of treatment. Chronic ethanol treatment resulted in a significant induction of ethylmorphine and benzphetamine N-demethylase activity in both LS and SS mice with the SS selected line showing a greater (P < .05) maximal induction. These data are suggestive that the induction of MEOS may have some significance in the acquisition of metabolic tolerance to ethanol and that there may be a genetic predisposition for these adaptive responses.

Paradoxical effects of d‐amphetamine upon seizure susceptibility in 2 selectively bred lines of mice

The ontogeny and substrates of amphetamine-induced changes in flurothyl-induced myoclonic and clonic seizure thresholds were investigated in 2 selectively bred lines of mice. The long-sleep mice exhibited dose-dependent increases in myoclonic and clonic susceptibility following amphetamine, irrespective of age. The noradrenergic agonist clonidine and the dopaminergic agonist apomorphine produced increases in susceptibility comparable to those seen with amphetamine. The short-sleep mice, however, exhibited a dichotomous myoclonic response to amphetamine that was age-dependent. Between 15 and 35 days of age amphetamine decreased seizure susceptibility whereas increases in susceptibility were noted at later ages. With the exception of 80 days, amphetamine did not affect clonic thresholds. In respect to myoclonus, clonidine persisted in producing effects similar to those seen in the long-sleep mice whereas apomorphine exhibited the same ontogenetic alteration in effect seen with amphetamine. These results confirm that the short-sleep mice might be a naturally occurring animal model of preadolescent hyperkinesis. Furthermore, the neuropharmacological tests demonstrate that the paradoxical response to amphetamine in the young short-sleep mice is mediated via a dopaminergic mechanism that must undergo dramatic change during ontogeny.

Maturational changes related to dopamine in the effects of d-amphetamine, cocaine, nicotine, and strychnine on seizure susceptibility.

The effects of four neural excitants (damphetamine, cocaine, nicotine, and strychnine) on myoclonic and clonic seizure susceptibility were investigated in two age groups (30 and 120 days) of short-sleep mice. Amphetamine and cocaine decreased susceptibility to myoclonus in young mice and increased susceptibility in mature mice. These effects were attenuated by pretreatment with haloperidol, indicating mediation by a dopaminergic system. Amphetamine did not alter clonic susceptibility in either age group of mice, whereas cocaine affected clonic susceptibility and myoclonus. These effects were not attenuated by haloperidol, indicating mediation by systems other than dopamine. Nicotine decreased susceptibility to myoclonus and increased susceptibility to clonus, whereas strychnine increased susceptibility to both types of seizure. Haloperidol, however, failed to alter any of these effects. These results are consistent with our previous work which suggests that a dopaminergic mechanism in these mice undergoes marked developmental changes between 30 and 120 days of age.

Electrophysiological responses to ethanol, pentobarbital, and nicotine in mice genetically selected for differential sensitivity to ethanol.

Cortical electroencephalographic (EEG) changes induced by ethanol (4.3 and 1.4 g/kg, ip), pentobarbital (50 and 16 mg/kg), and nicotine (1.0 g/kg) were examined in long-sleep (LS) and short-sleep (SS) mice that were genetically selected for differential sleep times induced by a hypnotic dosage of ethanol. Ethanol (4.3 g/kg) caused EEG changes that paralleled the behavioral differences, whereas no differences between selected lines were observed following the activating dose (1.4 g/kg). Data support the notion that the known difference in ethanol sleep times is due not to greater SS sensitivity to ethanol activation but rather to greater LS sensitivity to ethanol hypnosis. No differences between selected lines were observed following 50 mg/kg pentobarbital, which again parallels previous behavioral data. The SS mice were more responsive to pentobarbital activation (16 mg/kg). Nicotine more severely reduced EEG power and heart rate in LS mice; a continuous iv infusion of nicotine elicited a distinct pattern of behavioral stereotypy for each selected line, with more profound motor and reflex depression in LS mice. The lines do not differ in rate of nicotine metabolism, hence they must differ in central nervous system sensitivity to nicotine. Thus, lines of mice selectively bred for differential sensitivity to ethanol also display marked differences in electrophysiological and behavioral responses to nicotine.

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The time constant of the decay (τ) of miniature end-plate currents (MEPCs) recorded from muscles excised from long-sleep (LS) and short-sleep (SS) mice was measured as an indicator of the mean open time of the channel which is gated by the nicotinic acetylcholine receptor. Similar τ values were recorded from LS and SS mice, and these values were similarly increased by acute ethanol (50 mM). Mice were then chronically treated with ethanol to induce tolerance to its hypnotic action. The time course of the development of tolerance was similar in the LS and SS lines. While MEPCs recorded from tolerant mice yielded similar τ values to controls, acute exposure to ethanol did not cause an increase. It is suggested that an adaptation takes place in the nicotinic acetylcholine receptor channel complex during the induction of tolerance to ethanol.

Alcohol intake in selected lines of mice: Importance of sex and genotype.

Mice selected by McClearn and Kakihana for differences in ethanol-induced sleep time were used as subjects. In Experiment 1, mice from the long-sleep (LS) and short-sleep (SS) lines were offered a choice of water or solution GS consisting of 3% glucose and .16% sodium saccharin (w/v); or a choice of water or solution GS + E that contained GS solution plus 4% ethanol (w/v). In Experiment 2, mice from the first experiment were provided with a three-way choice among water, solution GS, and solution GS + E. In both experiments, SS mice (alcohol-insensitive) consumed more GS + E than LS mice (alcohol-sensitive). In addition, female mice drank considerably more GS + E solution than male mice. Thus, consumption of sweetened ethanol in both a two-way choice (water and GS + E) and a three-way choice (water, GS, and GS + E) is dependent on both genotype and sex. High genetic sensitivity to ethanol was associated with low consumption, and vice versa. Although females consumed more alcohol than males, famales of these lines have not been previously found to show lower sensitivity to acute alcohol administration.

Dissociation between the anticonvulsant action of alcohol and its depressant action in mice of different genotypes.

Abstract The effects of ethanol and pentobarbital on flurothyl seizure susceptibility were studied in two lines of mice that were derived by selective breeding with respect to ethanol sleep time. Short-sleep mice (SS) were more susceptible than long-sleep mice (LS) to flurothyl-induced myoclonus, but there was no line difference in susceptibility to clonus. Low doses of ethanol or pentobarbital increased seizure latencies in both populations, and there was no difference between the lines in the effect of these drugs. The lack of difference in response to ethanol in the two populations indicates that the genetic mechanisms which determine sensitivity to ethanol's anticonvulsant action are not identical with those which determine sensitivity to its depressant action.

Activating and anesthetic effects of general depressants

The long-sleep (LS) and short-sleep (SS) lines of mice were derived by selective breeding with respect to ethanol sleep time. We found that in current generations LS mice also have longer sleep times than SS mice to trichloroethanol and paraldehyde. Two subsequent experiments tested our hypothesis that mice that are relatively insensitive to the hypnotic effects of depressant drugs might be relatively activated by low doses of these drugs. Both experiments failed to support the hypothesis. First, although SS mice were more activated than LS mice by subhypnotic doses of paraldehyde, the lines did not differ in the degree of activation produced by low doses of trichloroethanol. Second, among mice from a genetically heterogeneous population (HS), there was no relation between the degree of activation induced by a low dose of ethanol and sensitivity to the hypnotic effects of a higher dose.

Behavioral effects of salsolinol and ethanol on mice selected for sensitivity to alcohol-induced sleep time

Two lines of mice, selected for differential sensitivity to alcohol, were tested following intracisternal administration of salsolinol, a compound putatively formed following ethanol ingestion. Both lines showed dose-dependent decreases in activity following salsolinol. When injected with ethanol, alcohol-insensitive short-sleep (SS) mice increased in activity while alcohol-sensitive long-sleep (LS) mice showed no change. Measures of co-ordination, taken following ethanol, demonstrated that LS mice were more incapacitated than SS mice. Salsolinol effects are discussed with reference to possible adrenergic blocking actions. Genetic differences in sensitivity to the incoordinating effects of alcohol were found.

Fertility and offspring survival in mice selected for different sensitivities to alcohol.

The LS (long-sleep) and SS (short-sleep) selected lines of mice exhibit relatively high and low sensitivity to alcohol, respectively, because of their previous history of selective breeding. The present study identifies other differences between the two lines. SS animals are almost twice as fertile as LS animals. Analysis of vaginal plug formation suggested that higher sexual activity of SS mice is an important component of their greater fertility. Forced maternal drinking of a 10% ethanol solution during gestation, and especially during lactation, caused a decrease in survival of LS but not of SS offspring. Increased cannibalism of pups by LS mothers was probably an important component of this decrease in progeny survival. Maternal consumption of liquid in both lines was decreased during gestation, and especially during lactation, when 10% ethanol was the only available liquid in each case. The physiological basis for decreased survival of LS progeny may involve either direct effects of alcohol on progeny or indirect effects caused by water, nutritional, or hormonal imbalance in LS mothers when they are exposed to alcohol stress. The relationship of fertility and progeny survival differences in the two lines to their differences in alcohol sensitivity produced by selective breeding is discussed.

Adrenocortical function in mice selectively bred for different sensitivity to ethanol

The adrenocortical response to ethanol, histamine and electric shock was studied in the Short-Sleep (SS) and Long-Sleep (LS) mice. The plasma corticosterone levels measured at 60 min following ethanol injection were consistently higher in LS mice (males and females) than in SS mice. The corticosterone levels determined 60 min after histamine injection (50 mg/kg) or electric foot shock were not statistically different in these two lines of mice. The initial rise of the corticosteroid after ethanol injection was identical in SS and LS mice, whereas the costicosteroid response to mild stress of saline injection was different: SS mice showed higher levels of corticosterone than LS mice at 20 and 30 min after the injection. Corticosterone increment attributable to alcohol stress clearly seemed greater in LS than in SS mice, even though the adrenals of the SS mice were 20–30% heavier than those of LS. The possible significance of these endocrine differences with respect to sensitivity to ethanol and behavioral arousal level was discussed.