Genetic analysis of the psychostimulant effects of nicotine in chromosome substitution strains and F2 crosses derived from A/J and C57BL/6J progenitors

Previous research utilizing the AcB/BcA recombinant congenic strains (RCS) of mice mapped provisional quantitative trait loci (QTLs) for the psychostimulant effects of nicotine to multiple regions on chromosomes 7, 11, 12, 14, 16, and 17. The current study was designed to confirm these QTLs in an A/J (A) × C57Bl/6J (B6) F2 cross and a panel of B6.A chromosome substitution strains (CSS). The panel of B6.A CSS consists of 21 strains, each carrying a different A/J chromosome on a B6 background. The A × B6 F2, CSS, A, and B6 mice were tested for sensitivity to the effects of nicotine on locomotor activity using a computerized open-field apparatus. In A × B6 F2 mice two QTLs were identified which confirm those previously observed in the AcB/BcA RCS. Significant differences in the expression of nicotine-induced activity were associated with loci on chromosome 11 (D11Mit62) and chromosome 16 (D16Mit131) in the A × B6 F2. At the chromosome 11 QTL, an A allele was associated with lower nicotine-induced activity scores relative to the B6. In contrast, the A allele was associated with greater relative nicotine activity values for the chromosome 16 QTL. A survey of the CSS panel confirmed the presence of QTLs for nicotine activation on chromosomes 2, 14, 16, and 17 previously identified in the AcB/BcA RCS. In the informative CSS strains, A alleles were consistently associated with greater nicotine-induced activity scores compared to the B6. The results of the present study are the first to validate QTLs for sensitivity to the effects of nicotine across multiple strains of mice. QTLs on chromosomes 2, 11, 14, 16, and 17 were confirmed in CSS and/or F2 mice. Significantly, the identification of a QTL on chromosome 16 has now been replicated in three crosses derived from the A and B6 progenitors.     

Recombinant congenic strains derived from A/J and C57BL/6J: a tool for genetic dissection of complex traits

Complex genetic traits can be dissected in mice, using well-defined sets of recombinant inbred strains, congenic strains, and recombinant congenic strains (RCS). We report the creation of a series of 37 independent RCS derived from the commonly used inbred strains of laboratory mouse A/J (A) and C57BL/6J (B6). These RCS were derived by systematic inbreeding of independent pairs of animals from a (F1 x A) x A and a (F1 x B) x B double backcross (N3), to create AcB and BcA strains, respectively. Fifteen AcB strains and 22 BcA strains at between 18 and 30 generations of inbreeding have been generated, are healthy, and show stable breeding performance. These strains have been genotyped for a total of 625 informative microsatellite DNA markers covering the entire genome, with an average spacing of 2.6 cM. Haplotype analyses indicate that on average, AcB and BcA strains contain 13.25% of the donor genome, a value close to the 12.5% expected from the breeding scheme used in their creation. In the AcB set, approximately 79% of the B6 genome has been transferred in independent strains, while in the BcA set approximately 84% of the A genome is represented on the B6 background. This represents an excellent coverage of congenic segments from both parental genomes in the two sets of strains, which can now be used to map simple and complex traits in a genome-wide fashion. As an example of the power of AcB/BcA strains as a mapping tool, the 37 strains were typed for susceptibility to infection with Legionella pneumophila, a monogenic trait controlled by the Lgn1 locus on Chromosome 13. Analysis of the strain distribution pattern of L. pneumophila susceptibility allowed direct mapping of Lgn1 to a 3-cM interval. The AcB/BcA set should prove a useful tool with which to investigate the complex genetic basis of known interstrain differences between A and B6 for many important diseases.     

Mapping of a Chromosome 12 Region Associated with Airway Hyperresponsiveness in a Recombinant Congenic Mouse Strain and Selection of Potential Candidate Genes by Expression and Sequence Variation Analyses

In a previous study we determined that BcA86 mice, a strain belonging to a panel of AcB/BcA recombinant congenic strains, have an airway responsiveness phenotype resembling mice from the airway hyperresponsive A/J strain. The majority of the BcA86 genome is however from the hyporesponsive C57BL/6J strain. The aim of this study was to identify candidate regions and genes associated with airway hyperresponsiveness (AHR) by quantitative trait locus (QTL) analysis using the BcA86 strain. Airway responsiveness of 205 F2 mice generated from backcrossing BcA86 strain to C57BL/6J strain was measured and used for QTL analysis to identify genomic regions in linkage with AHR. Consomic mice for the QTL containing chromosomes were phenotyped to study the contribution of each chromosome to lung responsiveness. Candidate genes within the QTL were selected based on expression differences in mRNA from whole lungs, and the presence of coding non-synonymous mutations that were predicted to have a functional effect by amino acid substitution prediction tools. One QTL for AHR was identified on Chromosome 12 with its 95% confidence interval ranging from 54.6 to 82.6 Mbp and a maximum LOD score of 5.11 (p = 3.68×10⁻³). We confirmed that the genotype of mouse Chromosome 12 is an important determinant of lung responsiveness using a Chromosome 12 substitution strain. Mice with an A/J Chromosome 12 on a C57BL/6J background have an AHR phenotype similar to hyperresponsive strains A/J and BcA86. Within the QTL, genes with deleterious coding variants, such as Foxa1, and genes with expression differences, such as Mettl21d and Snapc1, were selected as possible candidates for the AHR phenotype. Overall, through QTL analysis of a recombinant congenic strain, microarray analysis and coding variant analysis we identified Chromosome 12 and three potential candidate genes to be in linkage with airway responsiveness.     

Identification of loci controlling restriction of parasite growth in experimental Taenia crassiceps cysticercosis

Human neurocysticercosis (NC) caused by Taenia solium is a parasitic disease of the central nervous system that is endemic in many developing countries. In this study, a genetic approach using the murine intraperitoneal cysticercosis caused by the related cestode Taenia crassiceps was employed to identify host factors that regulate the establishment and proliferation of the parasite. A/J mice are permissive to T. crassiceps infection while C57BL/6J mice (B6) are comparatively restrictive, with a 10-fold difference in numbers of peritoneal cysticerci recovered 30 days after infection. The genetic basis of this inter-strain difference was explored using 34 AcB/BcA recombinant congenic strains derived from A/J and B6 progenitors, that were phenotyped for T. crassiceps replication. In agreement with their genetic background, most AcB strains (A/J-derived) were found to be permissive to infection while most BcA strains (B6-derived) were restrictive with the exception of a few discordant strains, together suggesting a possible simple genetic control. Initial haplotype association mapping using >1200 informative SNPs pointed to linkages on chromosomes 2 (proximal) and 6 as controlling parasite replication in the AcB/BcA panel. Additional linkage analysis by genome scan in informative [AcB55xDBA/2]F1 and F2 mice (derived from the discordant AcB55 strain), confirmed the effect of chromosome 2 on parasite replication, and further delineated a major locus (LOD = 4.76, p<0.01; peak marker D2Mit295, 29.7 Mb) that we designate Tccr1 (T. crassiceps cysticercosis restrictive locus 1). Resistance alleles at Tccr1 are derived from AcB55 and are inherited in a dominant fashion. Scrutiny of the minimal genetic interval reveals overlap of Tccr1 with other host resistance loci mapped to this region, most notably the defective Hc/C5 allele which segregates both in the AcB/BcA set and in the AcB55xDBA/2 cross. These results strongly suggest that the complement component 5 (C5) plays a critical role in early protective inflammatory response to infection with T. crassiceps.     

Genetic control of high density lipoprotein-cholesterol in AcB/BcA recombinant congenic strains of mice

Epidemiological studies show that high HDL-cholesterol (HDLc) decreases the risk of cardiovascular disease. To map genes controlling lipid metabolism, particularly HDLc levels, we screened the plasma lipids of 36 AcB/BcA RC mouse strains subjected to either a normal or a high-fat/cholesterol diet. Strains BcA68 and AcB65 showed deviant HDLc plasma levels compared with the parental A/J and C57BL/6J strains; they were thus selected to generate informative F2 crosses. Linkage analyses in the AcB65 strain identified a locus on chromosome 4 (Hdlq78) responsible for high post-high fat diet HDLc levels. This locus has been previously associated at genome-wide significance to two regions in the human genome. A second linkage analysis in strain BcA68 identified linkage in the vicinity of a gene cluster known to control HDLc levels. Sequence analysis of these candidates identified a de novo, loss-of-function mutation in the ApoA1 gene of BcA68 that prematurely truncates the ApoA1 protein. The possibility of dissecting the specific effects of this new ApoA1 deficiency in the context of isogenic controls makes the BcA68 mouse a valuable new tool.     

Genetic analysis of alcohol intake in recombinant inbred and congenic strains derived from A/J and C57BL/6J progenitors

The objective of the present study was to map quantitative trait loci (QTL) for alcohol intake using A × B/B × A recombinant inbred (RI) and AcB/BcA recombinant congenic (RC) strains of mice that were independently derived from the A/J and C57BL/6J progenitors. Mice were screened for levels of alcohol consumption with four days of forced exposure to alcohol, followed by three weeks of free choice between water and a 10% alcohol solution. Alcohol consumption data previously collected for 27 A × B/B × A RI strains were reanalyzed using a larger marker set and composite interval mapping. The reanalysis found markers on Chromosome 2 (D2Mit74, 107 cM) (males and females) and on Chromosome 11 (Pmv22, 8 cM) (females only) that exceeded the threshold for significant loci, and found suggestive loci (in males) on Chromosomes 10 (D10 Mit126, 21 cM), 12 (D12Mit37, 1 cM), 15 (Pdgfb, 46.8 cM), and 16 (D16Mit125, 29 cM). An additional suggestive locus was identified in female RI mice on Chromosome 11 (D11Mit120, 47.5 cM). Composite interval mapping (CIM) analysis indicated that there was a significant association between loci at Pdgfb and D2Mit74 in both males and females. Analysis of the AcB/BcA RC strains identified 11 QTL on Chromosomes 2, 3, 5,6, 7, 8, 9, 10, 12, 13, and 15. QTL on Chromosomes 7, 10, 12, and 15 were identified in both the A × B/B × A RI and AcB/BcA RC strains of mice. Additional QTLs identified on Chromosomes 2, 3, 7, 11, and 15 overlap with those previously identified in the literature using strains of mice with a C57BL/6J progenitor.     

Mapping of clinical and expression quantitative trait loci in a sex-dependent effect of host susceptibility to mouse-adapted influenza H3N2/HK/1/68

Seasonal influenza outbreaks and recurrent influenza pandemics present major challenges to public health. By studying immunological responses to influenza in different host species, it may be possible to discover common mechanisms of susceptibility in response to various influenza strains. This could lead to novel therapeutic targets with wide clinical application. Using a mouse-adapted strain of influenza (A/HK/1/68-MA20 [H3N2]), we produced a mouse model of severe influenza that reproduces the hallmark high viral load and overexpression of cytokines associated with susceptibility to severe influenza in humans. We mapped genetic determinants of the host response using a panel of 29 closely related mouse strains (AcB/BcA panel of recombinant congenic strains) created from influenza-susceptible A/J and influenza-resistant C57BL/6J (B6) mice. Combined clinical quantitative trait loci (QTL) and lung expression QTL mapping identified candidate genes for two sex-specific QTL on chromosomes 2 and 17. The former includes the previously described Hc gene, a deficit of which is associated with the susceptibility phenotype in females. The latter includes the phospholipase gene Pla2g7 and Tnfrsf21, a member of the TNFR superfamily. Confirmation of the gene underlying the chromosome 17 QTL may reveal new strategies for influenza treatment.     

A major QTL on chromosome 11 influences psychostimulant and opioid sensitivity in mice

The identification of genes influencing sensitivity to stimulants and opioids is important for determining their mechanism of action and may provide fundamental insights into the genetics of drug abuse. We used a panel of C57BL/6J (B6; recipient)× A/J (donor) chromosome substitution strains (CSSs) to identify quantitative trait loci (QTL) for both open field activity and sensitivity to the locomotor stimulant response to methamphetamine (MA). Mice were injected with saline (days 1 and 2) and MA (day 3; 2 mg/kg i.p.). We analyzed the total distance traveled in the open field for 30 min following each injection. CSS-8, -11 and -16 showed reduced MA-induced locomotor activity relative to B6, whereas CSS-10 and -12 showed increased MA-induced locomotor activity. Further analysis focused on CSS-11 because it was robustly different from B6 following MA injection, but did not differ in activity following saline injection and because it also showed reduced locomotor activity in response to the mu-opioid receptor agonist fentanyl (0.2 mg/kg i.p.). Thus, CSS-11 captures QTLs for the response to both psychostimulants and opioids. Using a B6 × CSS-11 F₂ intercross, we identified a dominant QTL for the MA response on chromosome 11. We used haplotype association mapping of cis expression QTLs and bioinformatic resources to parse among genes within the 95% confidence interval of the chromosome 11 QTL. Identification of the genes underlying QTLs for response to psychostimulants and opioids may provide insights about genetic factors that modulate sensitivity to drugs of abuse.     

Sensitivity to the locomotor-stimulant effects of ethanol and allopregnanolone: a quantitative trait locus study of common genetic influence

Previous studies have suggested that common genetic mechanisms influence sensitivity to the locomotor-stimulant effects of ethanol and allopregnanolone. We conducted two quantitative trait locus (QTL) studies to identify chromosomal regions that harbor genes that influence locomotor response to ethanol (2 g/kg) and allopregnanolone (17 mg/kg) using F2 crosses between C57BL/6J and DBA/2J mice. Because our previous data from the BXD recombinant inbred strains had indicated that chromosome 2 contained QTL for sensitivity to the locomotor-stimulant effects of both ethanol and allopregnanolone, we also tested reciprocal chromosome 2 congenic strains for sensitivity to the locomotor-stimulant effects of both drugs. The F2 analysis for ethanol sensitivity identified significant QTL on chromosomes 1 and 2 and suggestive QTL on chromosomes 5 and 9. The analysis of the allopregnanolone F2 study identified suggestive QTL on chromosomes 3, 5 and 12. Suggestive evidence for a female-specific QTL on chromosome 2 was also found. The studies of congenic mouse strains indicated that both the congenic strains captured one or more QTL for sensitivity to the locomotor-stimulant effects of both ethanol (2 g/kg) and allopregnanolone (17 mg/kg). When Fisher's method was used to combine the P values for the RI, F2 and congenic studies of the chromosome 2 QTL, cumulative probability scores of 9.6 x 10(-15) for ethanol and 7.7 x 10(-7) for allopregnanolone were obtained. These results confirm the presence of QTL for ethanol and allopregnanolone sensitivity in a common region of chromosome 2 and suggest possible pleiotropic genetic influence on sensitivity to these drugs.     

Nicotine-Induced Sensitization in Mice: Changes in Locomotor Activity and Mesencephalic Gene Expression

It is believed that drug-induced behavioral sensitization is an important process in the development of substance dependence. In order to explore mechanisms of sensitization, a mouse model of nicotine-induced locomotor sensitization was established, and effects of the sensitization process on mesencepahlic gene expression were examined. A schedule, which included 3 weeks of intermittent nicotine exposure (0.5 mg/kg, s.c.) and 3 weeks of withdrawal, resulted in locomotor sensitization. Effects of sensitization on mesencephalic expression of approximately 14,000 genes were assessed using oligonucleotide microarrays. Signal intensity differences in samples obtained from repeated nicotine- and saline-exposed animals were analyzed with z-test after False Discovery Rate (FDR) multiple test correction. Genes related to GABA-A receptors and protein phosphatases were among 68 genes showing significantly different expression levels between the saline and the nicotine groups. We hypothesize that some of the gene expression changes in the mesencephalon are involved in pathways leading to nicotine-induced sensitization. Down-regulation of GABA-A receptors induced by repeated nicotine exposure may facilitate dopaminergic neuronal transmission and may contribute to increased locomotor activity.     

Genetic analysis of the psychomotor stimulant effect of ethanol

Genetic influences on the psychomotor stimulant effect of ethanol may be a key feature of abuse liability. While earlier work has shown the activational effects of ethanol to be under the influence of a relatively uncomplicated additive genetic system, preliminary data from our laboratory suggested the possibility of nonadditive genetic variance. In the present study, a full Mendelian cross was conducted to further characterize gene action and search for quantitative trait loci (QTL) influencing the psychomotor stimulant properties of ethanol. We tested 3062 mice of the six Mendelian cross genotypes (P1, P2, F1, F2, BC1 and BC2) derived from a cross between the C57BL/6J (B6) and C3H/HeJ (C3H) inbred strains of mice. On day 1, mice were injected with saline, put in a holding cage for 5 min, then placed in an activity monitor for 5 min. On day 2, mice were injected with 1.5 g/kg ethanol, and activity again monitored for 5 min. Analysis showed the expected activation in the C3H strain and little activation in the B6 strain, with no effect of sex. Biometrical genetic analysis showed a best-fit model that included the mean (m), additive effect (a), and an epistatic parameter (i = homozygote by homozygote interaction). Analysis showed good evidence for QTL on chromosomes 1 (logarithm of odds (LOD) 3.4-7.5, 88-100 cM), 6 (LOD 9.1-10.4, 46-50 cM) and 15 (LOD 7.3-8.8, 28-32 cM). While the regions on chromosomes 1 and 6 have previously been implicated in several different ethanol-related phenotypes, this is the first report of a QTL influencing the psychomotor stimulant properties of ethanol on chromosome 15. Other studies have identified QTL in this region of chromosome 15 mediating locomotor activation caused by other psychostimulants, including cocaine, amphetamine and phencyclidine.     

Environmental Enrichment Alters Nicotine-Mediated Locomotor Sensitization and Phosphorylation of DARPP-32 and CREB in Rat Prefrontal Cortex

Exposure within an environmental enrichment paradigm results in neurobiological adaptations and decreases the baseline of locomotor activity. The current study determined activation of DARPP-32 (dopamine- and cAMP-regulated phosphoprotein-32) and CREB (cAMP response element binding protein), and locomotor activity in rats raised in enriched (EC), impoverished (IC), and standard (SC) conditions following repeated administration of nicotine or saline. In the saline-control group, the basal phosphorylation state of DARPP-32 at Threonine-34 site (pDARPP-32 Thr34) in the prefrontal cortex (PFC) was lower in EC compared to IC and SC rats, which was positively correlated with their respective baseline activities. While nicotine (0.35 mg/kg, freebase) produced locomotor sensitization across all housing conditions when the nicotine-mediated locomotor activity was expressed as a percent change from their respective saline control, EC rats displayed greater sensitization to nicotine than IC and SC rats. Consistent with the behavioral findings, repeated nicotine injection increased pDARPP-32 Thr34 in PFC of EC and IC rats and in nucleus accumbens of EC rats; however, the magnitude of change from saline control in nicotine-induced enhancement of pDARPP-32 Thr34 in PFC was strikingly increased in EC rats relative to IC rats. Moreover, EC rats had lower basal phosphorylation levels of CREB at serine 133 in PFC and nucleus accumbens compared to IC and SC rats, whereas the nicotine-induced increase in phosphorylated CREB-Ser133 was more pronounced in PFC of EC rats relative to IC and SC rats. Collectively, these findings suggest innovative insights into advancing our understanding of the molecular mechanisms of enrichment-induced changes in the motivational effects of nicotine, and aiding in the identification of new therapeutic strategies for tobacco smokers.     

WIN-55,212-2 and SR-141716A alter nicotine-induced changes in locomotor activity, but do not alter nicotine-evoked [3H]dopamine release

Nicotine, the main psychoactive ingredient in tobacco, plays a key role in the development of cigarette smoking addiction. The endocannabinoid system has been demonstrated to have an important role in the motivational and reinforcing effects of drugs. The present study used behavioral and neurochemical techniques to study the interaction of cannabinoid receptors and nicotine pharmacology. In a locomotor activity experiment in rats, the CB(1)/CB(2) cannabinoid receptor agonist WIN-55,212-2 (0.28-2.8 mg/kg) attenuated nicotine (0.4 mg/kg)-induced hyperactivity, but did not alter nicotine (1.0 mg/kg)-induced hypoactivity. In contrast, the selective CB(1) cannabinoid receptor antagonist SR-141716A (1.0 mg/kg) diminished nicotine-induced hypoactivity, but did not alter nicotine-induced hyperactivity. In a neurochemical experiment, rat striatal slices preloaded with [(3)H]dopamine were superfused with WIN-55,212-2 or SR-141716A. A high concentration (100 microM) of WIN-55,212-2 evoked [(3)H]overflow, but this effect was not blocked by the cannabinoid receptor antagonist AM-251. SR-141716A did not evoke [(3)H]overflow, and neither WIN-55,212-2 nor SR-141716A altered nicotine-evoked [(3)H]overflow. Overall, these results indicate a behavioral interaction between cannabinoid receptors and nicotine pharmacology. Likely, WIN-55,212-2 and SR-141716A block nicotine-induced changes in behavior through an indirect mechanism, such as alteration in endocannabinoid regulation of motor circuits, rather than directly through blockade of nicotinic acetylcholine receptors.     

Genetic control of susceptibility to carcinogen-induced colorectal cancer in mice: The Ccs3 and Ccs5 loci regulate different aspects of tumorigenesis

Colorectal cancer (CRC) is a multistep disease that involves a two-way interaction between a complex genetic pre-disposition component, and a set of poorly understood extrinsic environmental factors. In mice, CRC can be induced by treatment with azoxymethane (AOM). Using a set of AcB/BcA recombinant congenic strains derived from CRC-susceptible A/J and CRC-resistant C57Bl/6J (B6) progenitors, we previously detected the Ccs3 locus (colon cancer susceptibility locus 3) as a major regulator of CRC susceptibility. Phenotyping of additional AcB/BcA strains for susceptibility to AOM-induced CRC has refined the Ccs3 interval to a 6.7 Mb segment on chromosome 3. In addition, the presence of intermediate susceptibility phenotypes in individual AcB/BcA strains suggested additional gene effects regulating CRC susceptibility in A/J and B6 strains. Those were investigated by linkage analysis and whole genome scanning in a set of 208 informative (B6 x A/J)F2 progeny, using tumor multiplicity as a quantitative measure of susceptibility. This analysis validated the important role of Ccs3 in regulating this trait, and additionally detected contribution from a second locus on the distal portion of chromosome 9 (LOD = 3.76), that was given the temporary designation of Ccs5. Ccs5 modulates tumor multiplicity in F2 animals bearing at least one A/J-derived susceptibility allele at Ccs3, with A/J-derived Ccs5 susceptibility alleles being inherited in a recessive manner. There is a strong additive effect of Ccs3 and Ccs5 on tumor multiplicity in F2 mice: mice doubly homozygotes for A/J or B6 alleles at Ccs3 and Ccs5 show tumor numbers similar to those of parental A/J and B6, respectively. Interestingly, the Ccs5 region overlaps several quantitative trait loci previously reported to regulate intestinal homeostasis and susceptibility to intestinal colitis in mice and humans. Our findings identify a novel two-locus system regulating CRC susceptibility in mice, of which the relevance to human CRC can now be tested experimentally.     

Nicotine-induced sensitization to ambulatory stimulant effect produced by daily administration into the ventral tegmental area and the nucleus accumbens in rats.

Bilateral injections of nicotine (30 micrograms/side) into the ventral tegmental area (VTA) and the nucleus accumbens (NACC) increased the ambulatory activity in rats. Moreover, daily injections of nicotine (10, 20 and 30 micrograms/side) into the VTA and the NACC for 6 successive days produced sensitization to the ambulatory stimulant effect of nicotine. Sensitization produced by daily injections of nicotine (20 micrograms/side) into both the sites was maintained for withdrawal periods of 10 days. Mecamylamine (2 mg/kg, i.p.), SCH23390 (0.05 mg/kg, i.p.) and spiperone (0.1 mg/kg, i.p.) antagonized nicotine-induced sensitization to the ambulatory stimulant nicotine-induced sensitization to the ambulatory stimulant effect produced by daily injections into the VTA. These results suggest that nicotine-induced sensitization to the ambulatory stimulant effect involves the stimulation of the mesolimbic dopaminergic pathway through the nicotinic acetylcholine receptor (nAChR) in the VTA and the NACC.     

New quantitative trait loci for the genetic variance in circadian period of locomotor activity between inbred strains of mice

Provisional quantitative trait loci (QTL) for circadian locomotor period and wheel-running period have been identified in recombinant inbred (RI) mouse strains. To confirm those QTL and identify new ones, the genetic component of variance of the circadian period was partitioned among an F2 intercross of RI mouse strains (BXD19 and CXB07). First, a genomic survey using 108 SSLP markers with an average spacing of 15 cM was carried out in a population of 259 (BXD19 x CXB07)F2 animals. The genome-wide survey identified two significant QTL for period of locomotor activity measured by infrared photobeam crossings on mouse chromosomes 1 (lod score 5.66) and 14 (lod score 4.33). The QTL on distal chromosome 1 confirmed a previous report based on congenic B6.D2-Mtv7a/Ty mice. Lod scores greater than 2.0 were found on chromosomes 1, 2, 6, 12, 13, and 14. In a targeted extension study, additional genotyping was performed on these chromosomes in the full sample of 341 F2 progeny. The 6 chromosome-wide surveys identified 3 additional QTL on mouse chromosomes 6, 12, and 13. The QTL on chromosome 12 overlaps with circadian period QTL identified in several prior studies. For wheel-running period, the chromosome-wide surveys identified QTL on chromosomes 2 and 13 and one highly suggestive QTL on proximal chromosome 1. The results are compared to other published studies of QTL of circadian period.     

Multiple cross mapping (MCM) markedly improves the localization of a QTL for ethanol-induced activation

This study examines the use of multiple cross mapping (MCM) to reduce the interval for an ethanol response QTL on mouse chromosome 1. The phenotype is the acute locomotor response to a 1.5-g/kg i.p. dose of ethanol. The MCM panel consisted of the six unique intercrosses that can be obtained from the C57BL/6J (B6), DBA/2J (D2), BALB/cJ (C) and LP/J (LP) inbred mouse strains (N ≥ 600/cross). Ethanol response QTL were detected only with the B6xD2 and B6xC intercrosses. For both crosses, the D2 and C alleles were dominant and decreased ethanol response. The QTL information was used to develop an algorithm for sorting and editing the chromosome 1 Mit microsatellite marker set ( http://www.jax.org ) . This process yielded a cluster of markers between 82 and 85 cM (MGI). Evidence that the QTL was localized in or near this interval was obtained by the analysis of a sample ( n  = 550) of advanced cross heterogenous stock animals. In addition, it was observed that one of the BXD recombinant inbred strains (BXD-32) had a recombination in the interval of interest which produced the expected change in behavior. Overall, the data obtained suggest that the information available within existing genetic maps coupled with MCM data can be used to reduce the QTL interval. In addition, the MCM data set can be used to interrogate gene expression data to estimate which polymorphisms within the interval of interest are relevant to the QTL.     

Genetic differences in nicotine-induced conditioned taste aversion

Genetic differences in nicotine-induced conditioned taste aversion were examined using inbred mice. Adult male C57BL/6J, DBA/2J, BALB/cJ and C3H/heJ mice were adapted to a 2-h per day water access regimen. Subsequently, mice received nicotine injections (0.5, 1.0 or 2.0 mg/kg) immediately after 1-h access to a NaCl flavored solution. DBA and C3H mice developed dosedependent aversions to the nicotine-paired flavor. BALB mice showed only minor reductions in intake with no difference between the nicotine dose groups. C57BL mice did not show development of nicotine-induced conditioned taste aversion. These results demonstrate that nicotine's aversive motivational effect is strongly influenced by genotype. Further, genetic sensitivity (DBA mice) or insensitivity (C57BL mice) to nicotine-induced conditioned taste aversion was similar to reports of genetic sensitivity to ethanol's aversive effect measured in this design.     

Nicotine induced seizures blocked by mecamylamine and its stereoisomers.

Recent genetic research has shown that certain forms of epilepsy may arise from mutations in the genes encoding for the alpha7 and alpha4 neuronal nicotinic acetylcholine receptor (nAChR) ion channels. These receptors are also involved with the induction of nicotine-induced seizures. (+/-)-Mecamylamine (Inversine), a classic nAChR antagonist, potently inhibits nicotine-induced seizures. The purpose of the present study was to assess the inhibitory effects of (+/-)-mecamylamine and its stereoisomers on nicotine-induced seizures in male Sprague-Dawley rats. Rats received saline, (+/-)-mecamylamine, R-(-)-mecamylamine, or S-(+)-mecamylamine (s.c.) at doses of 0.1, 0.3, or 1.0 mg/kg 15 minutes prior to nicotine injection, 3.6 mg/kg (s.c.), an optimal dose for seizure induction. Rats were observed for 30 minutes with seizure latency, duration, and severity as primary measures and locomotor activity recorded as a secondary measure at 5-minute intervals. The results indicate that mecamylamine and each of its stereoisomers block nicotine-induced seizures in a dose-related manner and suggest that the S-(+/-)- mecamylamine isomer has inhibitory properties more similar to the racemic than to the R-(-)-mecamylamine isomer. The results of this study may be clinically important for the future design of novel anti-seizure medications.     

Differences in Nicotine-Induced Dopamine Release and Nicotine Pharmacokinetics Between Lewis and Fischer 344 Rats

Studies have shown a greater preference for the self-administration of drugs such as nicotine and cocaine in the Lewis rat strain than in the Fischer 344 strain. We examined some factors that could contribute to such a difference. The baseline level of extracellular dopamine in nucleus accumbens shell was about 3-times higher in Fischer rats than in Lewis rats (3.18 ± 0.26 vs. 1.09 ± 0.14 pg/sample). Nicotine (50-100 g/kg)-induced release of dopamine, expressed in absolute terms, was similar in the two strains. Dopamine release expressed in relative terms (as percent of baseline), however, was significantly greater in Lewis rats than in Fischer rats at 30 min after the first nicotine injection. We suggest that the relative increase is of more influence than the absolute level for determining preference; a lower physiological extracellular dopamine level thus represent a risk factor for increased preference. Amphetamine-induced dopamine release expressed in relative terms was not greater in the Lewis strain. In the initial time period of the microdialysis experiments, a sharper peak in nicotine-induced accumbal dopamine release in Lewis and a less but more sustained release in Fischer rats was observed. This release pattern paralleled the faster clearance of nicotine from blood of Lewis compared to Fischer rats. In tissue slices the electrically induced dopamine release was highest in the nucleus accumbens and lowest in the ventral tegmentum. A significant effect of nicotine was lowering the electrically induced release of dopamine in frontal cortex slices from Fischer brain and increasing this dopamine release in the ventral tegmentum of Lewis brain slices indicating that the ventral tegmentum, an area controlling dopamine release in the accumbens, is more responsive to nicotine in the Lewis rat. Nicotine levels tended to be more sustained in Fischer rats in different brain regions, although the difference in nicotine levels between the strains was not significant at any time period. Several factors contribute to nicotine preference, including the endogenous dopamine level, and the sensitivity of ventral tegmentum neurons to nicotine-induced dopamine release. Strain differences in pharmacokinetics of nicotine may also play a role.