Potential regulation of nicotine and ethanol actions by alpha4-containing nicotinic receptors.

A restriction fragment length polymorphism (RFLP) associated with a major nicotinic receptor subunit (i.e., alpha4) has been identified in two mouse lines that were selectively bred for differences in sensitivity to ethanol. These mice, referred to as Long-Sleep (LS) and Short-Sleep (SS) mice, also differ in sensitivity to several effects of nicotine. The potential role of the alpha4 RFLP in regulating several responses to nicotine and ethanol was evaluated by using the LSxSS-derived recombinant inbred (RI) strains. Those RI strains that carried the LS-like alpha4 RFLP were more sensitive to the depressant effects of nicotine on Y-maze crossing and rearing activities and ethanol-induced increases in Y-maze crossing activity than were those RI strains that carry the SS-like alpha4 RFLP. The LS-like RI strains were also more sensitive to nicotine-induced hypothermia. The RFLP was not associated with strain differences in ethanol-induced body temperature or sleep time. The potential role of the RFLP in regulating ethanol and nicotine consumption was evaluated in heterogeneous stock (HS) mice. An association was found between the alpha4 RFLP and variation in ethanol consumption, but not in nicotine consumption, as measured in a four-bottle choice test. Recent studies of ethanol and tobacco abuse by human beings suggest that common genes may influence these two forms of substance abuse. The results of the studies reported here suggest that the alpha4 nicotinic receptor gene should be evaluated for its potential role in regulating ethanol and tobacco abuse in human beings.

The role of RNA editing of kainate receptors in synaptic plasticity and seizures.

The ionotropic glutamate receptor subunit GluR6 undergoes developmentally and regionally regulated Q/R site RNA editing that reduces the calcium permeability of GluR6-containing kainate receptors. To investigate the functional significance of this editing in vivo, we engineered mice deficient in GluR6 Q/R site editing. In these mutant mice but not in wild types, NMDA receptor-independent long-term potentiation (LTP) could be induced at the medial perforant path-dentate gyrus synapse. This indicates that kainate receptors with unedited GluR6 subunits can mediate LTP. Behavioral analyses revealed no differences from wild types, but mutant mice were more vulnerable to kainate-induced seizures. Together, these results suggest that GluR6 Q/R site RNA editing may modulate synaptic plasticity and seizure vulnerability.

Mice lacking PKC gamma exhibit decreased anxiety.

To investigate the contribution of the PKC gamma isoform of protein kinase C (PKC) in neurochemical pathways regulating anxiety, mice lacking the gene encoding PKC gamma were tested with heterozygote and wild-type littermates in three approach-avoidance tests of anxiety. Null mutant mice consistently displayed a decrease in baseline anxiety-related behaviors in the elevated plus-maze, the black/white box, and the mirrored chamber. In the elevated plus-maze, mutant mice entered the open arms significantly more often and spent more time in the open arms of the maze. In the black/white box, transitions between the compartments were greatest in the null mutant mice, and in the mirrored chamber, mutant mice were markedly less anxious with significantly decreased latencies to enter and more time spent in the chamber. Indices of locomotor activity in the mazes and tests of activity in home cages indicated that the reduced anxiety observed in the mutant mice was not due to baseline locomotor activity differences among the genotypes. These results suggest that PKC gamma be considered as one factor in the etiology of anxiety, perhaps via its post-synaptic regulation of GABAA and 5-HT2 receptors, two receptors implicated in the neurobiology of anxiety.

Genetic and pharmacological strategies identify a behavioral function of neuronal nicotinic receptors.

The studies outlined here used pharmacological and genetic approaches to attempt to identify the nicotinic receptors that modulate nicotine-induced seizures. Full-blown clonic-tonic seizures were induced by intracerebroventricular (i.c.v.) injection of nicotine, the alpha4beta2 selective agonist ABT-418 and the alpha7-selective GTS-21. Cytisine, which is a partial agonist at alpha4beta2-type receptors, produced partial seizures. DHbetaE and MLA did not block nicotine-induced seizures. Instead, both antagonists caused seizures. Restriction fragment length polymorphisms (RFLPs) for the alpha7 receptor were identified in two inbred strains (C3H and DBA) that differ in sensitivity to nicotine-induced seizures. F2 mice derived from a C3H x DBA cross that were homozygous for the C3H variant of the alpha7 RFLP were more sensitive to nicotine-induced seizures than were F2 mice that were homozygous for the DBA RFLP. In a study that used RI strains derived from two selectively bred mouse lines (LS and SS), an association between sensitivity to nicotine-induced seizures and an RFLP associated with the alpha4 gene was found. These data support the assertion that both alpha4 and alpha7 receptor types are involved in modulating convulsions produced by nicotine.

Potential role of the alpha4 and alpha6 nicotinic receptor subunits in regulating nicotine-induced seizures.

Several studies have shown that genetic factors influence sensitivity to nicotine-induced seizures in the mouse. We used recombinant inbred (RI) strains derived from the Long-Sleep (LS) and Short-Sleep (SS) mouse lines to assess the possibility that polymorphisms associated with one or more of the nicotinic receptors cosegregate with differential sensitivity to nicotine-induced seizures. Restriction fragment length polymorphisms (RFLPs) associated with the alpha2, alpha3, alpha4, alpha5, and alpha6 nicotinic receptors were identified in the LS and SS mouse lines, but the RI strains were polymorphic for only the alpha4 and alpha6 RFLPs. The RI strains were tested for sensitivity to nicotine-induced seizures. Strain and gender effects on seizure sensitivity were obtained as assessed by ED(50) values and latency to seizure. Those RI strains with the LS-like alpha4 RFLP were, on average, more sensitive to nicotine-induced seizures than were those RI strains with SS-like alpha4 RFLP. The alpha6 nicotine receptor may also play a role in modulating nicotine-induced seizures, but this effect is markedly influenced by gender. Females of the RI strains with the LS-like alpha6 RFLP were more sensitive to nicotine than were females of the strains with the SS-like alpha6 RFLP. Similar trends were seen in the males, but these trends were not significant. Thus, these strain differences may be due to polymorphisms associated with both the alpha4 and alpha6 nicotinic receptors, but gender also plays an important role in regulating sensitivity to nicotine-induced seizure.

Classical and neoclassical approaches to the genetic analysis of alcohol-related phenotypes.

Theoretical descriptions are given for two breeding methods in animal genetics that might be of use in alcohol research. These methods are marker-based selection and marker-based development of congenic strains, both using DNA markers such as polymerase chain reaction-detectable polymorphisms as the criteria for breeding. Such designs would utilize these markers as indicators of adjacent Quantitative Trait Loci (QTL) that are influential on alcohol-related phenotypes. Issues in the logic and implementation of these methods, such as proximity of the markers and the QTL allele, are explored. A third method, development of congenic strains with phenotypic screening, is also described. This method is currently being used to create two sets of congenic lines on a C57BL/6 inbred mouse background. The criterion phenotype is locomotor activation to 1.5 g/kg (i.p.) ethanol. Data are reported on the success of transferring the activation phenotype from two strains, DBA/2Abg and MOLD/Rk-Abg, onto the nonactivated C57BL/6Abg background. The value of these methods in alcohol research is outlined with regard to both identification of relevant genes and for their use as tools in basic research on mechanism of alcohol action.

Biometrical genetic analysis of ethanol's psychomotor stimulant effect.

Hereditary influences on psychomotor stimulant effects of ethanol (ETOH) were studied in a classical Mendelian cross of DBA/2Abg (D2) and C57BL/6Abg (B6) inbred mouse strains. A dose-response study with nine doses (0-3.5 g/kg ip) indicated that B6 mice lack the activational limb of the biphasic curve (< or = 1.5 g/kg), as assessed in a 15-min test. D2 mice ran greater distances and ran faster, at doses up to 2.5 g/kg. B6 mice showed no increments in speed or distance at these doses that activated D2 mice. Several other indices reinforced the conclusion of ETOH-induced behavioral activation in D2 mice and lack there of in B6 mice (traditional photobeam interruptions--horizontal counts; center distance; vertical movements; inactive time; as well as derived indices of running speed and average length of each movement). The F1, F2, and backcross generations produced dose-response curves that showed additive inheritance of the activational response to doses below 1.5 g/kg. A second study (n = 1446) examined response to 1.5 g/kg with a within-subjects design in the full Mendelian cross. This study verified the completely additive mode of inheritance for the total distance measure suggested in the dose-response study, and showed that sex linkage and sex differences were not present. Narrow sense heritability of the ETOH activation response (indexed by total distance) was calculated at 0.35 and approximately 3 loci were estimated to be responsible for the B6/D2 difference. The other phenotypes (described above) also showed strongly additive genetic control.(ABSTRACT TRUNCATED AT 250 WORDS)

Genetic influences on locomotor activating effects of ethanol and sodium pentobarbital.

The paradoxical capability of sedative-hypnotics to produce behavioral disinhibition varies among genotypes. In DBA/2 mice ethanol (ETOH) produced strong locomotor stimulation with the peak of the biphasic curve at 1.5 g/kg IP. C57BL/6 mice showed no activation, and F1S were intermediate. These characterizations held for a variety of behavioral indices derived from 15 min tests, such as distance, speed, and rest time, at doses in the 0-2.0 g/kg range. Analogous studies with sodium pentobarbital (0-40 mg/kg) yielded a similar pattern of strain differences in locomotor stimulation. In contrast, loss of righting reflex durations (60 mg/kg PENTO, IP) were similar in the two strains, indicating dissociation of activating and sedative effects. In complementary studies, long- and short-sleep mice, which were bred for differences in soporific effects of ETOH, showed similar activation profiles at ETOH doses up to 1.5 g/kg and PENTO doses up to 30 mg/kg. These studies provide support for an hypothesis of common genetic control of the activation effect for ETOH and PENTO.

Differential activating effects of ethanol in C57BL/6Abg and DBA/2Abg mice.

A characteristic pattern of ETOH-induced locomotor activation in the DBA/2Abg strain (D2), small activation or sedation in the C57BL/6Abg (B6), and an intermediate position of the F1s was found using a between-group design and 1.5 g/kg ETOH. This pattern was consistent for a variety of behavioral indices not previously examined, including distance, rest time, movement speed and length, as well as the traditional horizontal counts. Using a within-subject, multiple day, repeated-testing procedure, the same three genotypes were also assessed after manipulating drug administration order, where ETOH exposure (1.5 g/kg) was on either the first or second test day. Another experiment examined the effect of lighting level on the response to 1.5 g/kg ETOH using a within-subjects approach. Neither the testing order nor lighting condition had any major influence on the magnitude of activation as measured by locomotor activity (distance). The pattern of additive genotypic influences exhibited by the B6, D2, and F1 mice is remarkably resistant to these contextual and procedural manipulations.