Polygenic contributions to performance on the Balloon Analogue Risk Task.

Risky decision-making is a common, heritable endophenotype seen across many psychiatric disorders. Its underlying genetic architecture is incompletely explored. We examined behavior in the Balloon Analogue Risk Task (BART), which tests risky decision-making, in two independent samples of European ancestry. One sample (n = 1138) comprised healthy participants and some psychiatric patients (53 schizophrenia, 42 bipolar disorder, 47 ADHD); the other (n = 911) excluded for recent treatment of various psychiatric disorders but not ADHD. Participants provided DNA and performed the BART, indexed by mean adjusted pumps. We constructed a polygenic risk score (PRS) for discovery in each dataset and tested it in the other as replication. Subsequently, a genome-wide MEGA-analysis, combining both samples, tested genetic correlation with risk-taking self-report in the UK Biobank sample and psychiatric phenotypes characterized by risk-taking (ADHD, Bipolar Disorder, Alcohol Use Disorder, prior cannabis use) in the Psychiatric Genomics Consortium. The PRS for BART performance in one dataset predicted task performance in the replication sample (r = 0.13, p = 0.000012, pFDR = 0.000052), as did the reciprocal analysis (r = 0.09, p = 0.0083, pFDR=0.04). Excluding participants with psychiatric diagnoses produced similar results. The MEGA-GWAS identified a single SNP (rs12023073; p = 3.24 × 10-8) near IGSF21, a protein involved in inhibitory brain synapses; replication samples are needed to validate this result. A PRS for self-reported cannabis use (p = 0.00047, pFDR = 0.0053), but not self-reported risk-taking or psychiatric disorder status, predicted behavior on the BART in our MEGA-GWAS sample. The findings reveal polygenic architecture of risky decision-making as measured by the BART and highlight its overlap with cannabis use.

Evaluation of delay discounting as a transdiagnostic research domain criteria indicator in 1388 general community adults.

BACKGROUND: The Research Domain Criteria (RDoC) approach proposes a novel psychiatric nosology using transdiagnostic dimensional mechanistic constructs. One candidate RDoC indicator is delay discounting (DD), a behavioral economic measure of impulsivity, based predominantly on studies examining DD and individual conditions. The current study sought to evaluate the transdiagnostic significance of DD in relation to several psychiatric conditions concurrently. METHODS: Participants were 1388 community adults (18-65) who completed an in-person assessment, including measures of DD, substance use, depression, anxiety, posttraumatic stress disorder, and attention-deficit hyperactivity disorder (ADHD). Relations between DD and psychopathology were examined with three strategies: first, examining differences by individual condition using clinical cut-offs; second, examining DD in relation to latent psychopathology variables via principal components analysis (PCA); and third, examining DD and all psychopathology simultaneously via structural equation modeling (SEM). RESULTS: Individual analyses revealed elevations in DD were present in participants screening positive for multiple substance use disorders (tobacco, cannabis, and drug use disorder), ADHD, major depressive disorder (MDD), and an anxiety disorder (ps < 0.05-0.001). The PCA produced two latent components (substance involvement v. the other mental health indicators) and DD was significantly associated with both (ps < 0.001). In the SEM, unique significant positive associations were observed between the DD latent variable and tobacco, cannabis, and MDD (ps < 0.05-0.001). CONCLUSIONS: These results provide some support for DD as a transdiagnostic indicator, but also suggest that studies of individual syndromes may include confounding via comorbidities. Further systematic investigation of DD as an RDoC indicator is warranted.

Identification of a novel, fast-acting GABAergic antidepressant.

Current pharmacotherapies for depression exhibit slow onset, side effects and limited efficacy. Therefore, identification of novel fast-onset antidepressants is desirable. GLO1 is a ubiquitous cellular enzyme responsible for the detoxification of the glycolytic byproduct methylglyoxal (MG). We have previously shown that MG is a competitive partial agonist at GABA-A receptors. We examined the effects of genetic and pharmacological inhibition of GLO1 in two antidepressant assay models: the tail suspension test (TST) and the forced swim test (FST). We also examined the effects of GLO1 inhibition in three models of antidepressant onset: the chronic FST (cFST), chronic mild stress (CMS) paradigm and olfactory bulbectomy (OBX). Genetic knockdown of Glo1 or pharmacological inhibition using two structurally distinct GLO1 inhibitors (S-bromobenzylglutathione cyclopentyl diester (pBBG) or methyl-gerfelin (MeGFN)) reduced immobility in the TST and acute FST. Both GLO1 inhibitors also reduced immobility in the cFST after 5 days of treatment. In contrast, the serotonin reuptake inhibitor fluoxetine (FLX) reduced immobility after 14, but not 5 days of treatment. Furthermore, 5 days of treatment with either GLO1 inhibitor blocked the depression-like effects induced by CMS on the FST and coat state, and attenuated OBX-induced locomotor hyperactivity. Finally, 5 days of treatment with a GLO1 inhibitor (pBBG), but not FLX, induced molecular markers of the antidepressant response including brain-derived neurotrophic factor (BDNF) induction and increased phosphorylated cyclic-AMP response-binding protein (pCREB) to CREB ratio in the hippocampus and medial prefrontal cortex (mPFC). Our findings indicate that GLO1 inhibitors may provide a novel and fast-acting pharmacotherapy for depression.

Fine-mapping of genes determining extrafusal fiber properties in murine soleus muscle.

Muscle fiber cross-sectional area (CSA) and proportion of different fiber types are important determinants of muscle function and overall metabolism. Genetic variation plays a substantial role in phenotypic variation of these traits; however, the underlying genes remain poorly understood. This study aimed to map quantitative trait loci (QTL) affecting differences in soleus muscle fiber traits between the LG/J and SM/J mouse strains. Fiber number, CSA, and proportion of oxidative type I fibers were assessed in the soleus of 334 genotyped female and male mice of the F34 generation of advanced intercross lines (AIL) derived from the LG/J and SM/J strains. To increase the QTL detection power, these data were combined with 94 soleus samples from the F2 intercross of the same strains. Transcriptome of the soleus muscle of LG/J and SM/J females was analyzed by microarray. Genome-wide association analysis mapped four QTL (genome-wide P < 0.05) affecting the properties of muscle fibers to chromosome 2, 3, 4, and 11. A 1.5-LOD QTL support interval ranged between 2.36 and 4.67 Mb. On the basis of the genomic sequence information and functional and transcriptome data, we identified candidate genes for each of these QTL. The combination of analyses in F2 and F34 AIL populations with transcriptome and genomic sequence data in the parental strains is an effective strategy for refining QTL and nomination of the candidate genes.

Discovery and refinement of muscle weight QTLs in B6 × D2 advanced intercross mice.

The genes underlying variation in skeletal muscle mass are poorly understood. Although many quantitative trait loci (QTLs) have been mapped in crosses of mouse strains, the limited resolution inherent in these conventional studies has made it difficult to reliably pinpoint the causal genetic variants. The accumulated recombination events in an advanced intercross line (AIL), in which mice from two inbred strains are mated at random for several generations, can improve mapping resolution. We demonstrate these advancements in mapping QTLs for hindlimb muscle weights in an AIL (n = 832) of the C57BL/6J (B6) and DBA/2J (D2) strains, generations F8-F13. We mapped muscle weight QTLs using the high-density MegaMUGA SNP panel. The QTLs highlight the shared genetic architecture of four hindlimb muscles and suggest that the genetic contributions to muscle variation are substantially different in males and females, at least in the B6D2 lineage. Out of the 15 muscle weight QTLs identified in the AIL, nine overlapped the genomic regions discovered in an earlier B6D2 F2 intercross. Mapping resolution, however, was substantially improved in our study to a median QTL interval of 12.5 Mb. Subsequent sequence analysis of the QTL regions revealed 20 genes with nonsense or potentially damaging missense mutations. Further refinement of the muscle weight QTLs using additional functional information, such as gene expression differences between alleles, will be important for discerning the causal genes.

Fine-mapping of muscle weight QTL in LG/J and SM/J intercrosses.

Genetic variation plays a substantial role in variation in strength, but the underlying mechanisms remain poorly understood. The objective of the present study was to examine the mechanisms underlying variation in muscle mass, a predictor of strength, between LG/J and SM/J strains, which are the inbred progeny of mice selected, respectively, for high and low body weight. We measured weight of five hindlimb muscles in LG/J and SM/J males and females, in F(1) and F(2) intercrosses, and in an advanced intercross (AI), F(34), between the two. F(2) mice were genotyped using 162 SNPs throughout the genome; F(34) mice were genotyped at 3,015 SNPs. A twofold difference in muscle mass between the LG/J and SM/J mouse strains was observed. Integrated genome-wide association analysis in the combined population of F(2) and AI identified 22 quantitative trait loci (QTL; genome-wide P < 0.05) affecting muscle weight on Chr 2 (2 QTL), 4, 5, 6 (7 QTL), 7 (4 QTL), 8 (4 QTL), and 11 (3 QTL). The LG/J allele conferred greater muscle weight in all cases. The 1.5-LOD QTL support intervals ranged between 0.3 and 13.4 Mb (median 3.7 Mb) restricting the list of candidates to between 5 and 97 genes. Selection for body weight segregated the alleles affecting skeletal muscle, the most abundant tissue in the body. Combination of analyses in an F(2) and AI was an effective strategy to detect and refine the QTL in a genome-wide manner. The achieved resolution facilitates further elucidation of the underlying genetic mechanisms affecting muscle mass.

The genetics of human personality.

Personality traits are the relatively enduring patterns of thoughts, feelings and behaviors that reflect the tendency to respond in certain ways under certain circumstances. Twin and family studies have showed that personality traits are moderately heritable, and can predict various lifetime outcomes, including psychopathology. The Research Domain Criteria characterizes psychiatric diseases as extremes of normal tendencies, including specific personality traits. This implies that heritable variation in personality traits, such as neuroticism, would share a common genetic basis with psychiatric diseases, such as major depressive disorder. Despite considerable efforts over the past several decades, the genetic variants that influence personality are only beginning to be identified. We review these recent and increasingly rapid developments, which focus on the assessment of personality via several commonly used personality questionnaires in healthy human subjects. Study designs covered include twin, linkage, candidate gene association studies, genome-wide association studies and polygenic analyses. Findings from genetic studies of personality have furthered our understanding about the genetic etiology of personality, which, like neuropsychiatric diseases themselves, is highly polygenic. Polygenic analyses have showed genetic correlations between personality and psychopathology, confirming that genetic studies of personality can help to elucidate the etiology of several neuropsychiatric diseases.

Heterogeneous stock rats: a model to study the genetics of despair-like behavior in adolescence.

Major depressive disorder (MDD) is a complex illness caused by both genetic and environmental factors. Antidepressant resistance also has a genetic component. To date, however, very few genes have been identified for major depression or antidepressant resistance. In this study, we investigated whether outbred heterogeneous stock (HS) rats would be a suitable model to uncover the genetics of depression and its connection to antidepressant resistance. The Wistar Kyoto (WKY) rat, one of the eight founders of the HS, is a recognized animal model of juvenile depression and is resistant to fluoxetine antidepressant treatment. We therefore hypothesized that adolescent HS rats would exhibit variation in both despair-like behavior and response to fluoxetine treatment. We assessed heritability of despair-like behavior and response to sub-acute fluoxetine using a modified forced swim test (FST) in 4-week-old HS rats. We also tested whether blood transcript levels previously identified as depression biomarkers in adolescent human subjects are differentially expressed in HS rats with high vs. low FST immobility. We demonstrate heritability of despair-like behavior in 4-week-old HS rats and show that many HS rats are resistant to fluoxetine treatment. In addition, blood transcript levels of Amfr, Cdr2 and Kiaa1539, genes previously identified in human adolescents with MDD, are differentially expressed between HS rats with high vs. low immobility. These data demonstrate that FST despair-like behavior will be amenable to genetic fine-mapping in adolescent HS rats. The overlap between human and HS blood biomarkers suggest that these studies may translate to depression in humans.

Selection for contextual fear conditioning affects anxiety-like behaviors and gene expression.

Conditioned fear and anxiety-like behaviors have many similarities at the neuroanatomical and pharmacological levels, but their genetic relationship is less well defined. We used short-term selection for contextual fear conditioning (FC) to produce outbred mouse lines with robust genetic differences in FC. The high and low selected lines showed differences in fear learning that were stable across various training parameters and were not secondary to differences in sensitivity to the unconditioned stimulus (foot shock). They also showed a divergence in fear potentiated startle, indicating that differences induced by selection generalized to another measure of fear learning. However, there were no differences in performance in a Pavlovian approach conditioning task or the Morris water maze, indicating no change in general learning ability. The high fear learning line showed greater anxiety-like behavior in the open field and zero maze, confirming a genetic relationship between FC and anxiety-like behavior. Gene expression analysis of the amygdala and hippocampus identified genes that were differentially expressed between the two lines. Quantitative trait locus (QTL) analysis identified several chromosomal regions that may underlie the behavioral response to selection; cis-acting expression QTL were identified in some of these regions, possibly identifying genes that underlie these behavioral QTL. These studies support the validity of a broad genetic construct that includes both learned fear and anxiety and provides a basis for further studies aimed at gene identification.

Acute and chronic responses to the convulsant pilocarpine in DBA/2J and A/J mice.

Characterizing the responses of different mouse strains to experimentally-induced seizures can provide clues to the genes that are responsible for seizure susceptibility, and factors that contribute to epilepsy. This approach is optimal when sequenced mouse strains are available. Therefore, we compared two sequenced strains, DBA/2J (DBA) and A/J. These strains were compared using the chemoconvulsant pilocarpine, because pilocarpine induces status epilepticus, a state of severe, prolonged seizures. In addition, pilocarpine-induced status is followed by changes in the brain that are associated with the pathophysiology of temporal lobe epilepsy (TLE). Therefore, pilocarpine can be used to address susceptibility to severe seizures, as well as genes that could be relevant to TLE. A/J mice had a higher incidence of status, but a longer latency to status than DBA mice. DBA mice exhibited more hippocampal pyramidal cell damage. DBA mice developed more ectopic granule cells in the hilus, a result of aberrant migration of granule cells born after status. DBA mice experienced sudden death in the weeks following status, while A/J mice exhibited the most sudden death in the initial hour after pilocarpine administration. The results support previous studies of strain differences based on responses to convulsants. They suggest caution in studies of seizure susceptibility that are based only on incidence or latency. In addition, the results provide new insight into the strain-specific characteristics of DBA and A/J mice. A/J mice provide a potential resource to examine the progression to status. The DBA mouse may be valuable to clarify genes regulating other seizure-associated phenomena, such as seizure-induced neurogenesis and sudden death.

Cdh13 and AdipoQ gene knockout alter instrumental and Pavlovian drug conditioning.

Genome-wide association studies in humans have suggested that variants of the cadherin-13 (CDH13) gene are associated with substance use disorder, subjective response to amphetamine, and attention deficit hyperactivity disorder. To examine the role of the Cdh13 and its peptide ligand adiponectin (AdipoQ) in addiction-related behaviors, we assessed Cdh13 knockout (KO) rats and AdipoQ KO mice using intravenous cocaine self-administration and conditioned place preference (CPP) paradigms. During intravenous cocaine self-administration, male Cdh13 heterozygous (+/-) and KO (-/-) rats showed increased cue-induced reinstatement compared with wild-type (WT) rats when presented with a cocaine-paired stimulus, whereas female Cdh13 rats showed no differences across genotype. Cdh13 -/- rats showed higher responding for a saccharin reinforcer and learned the choice reaction time (RT) task more slowly than WTs. However, we found no differences between Cdh13 -/- and +/+ rats in responding for sensory reinforcement, number of premature responses in the RT task, tendency to approach a Pavlovian food cue, CPP and locomotor activation to cocaine (10 or 20 mg/kg). In AdipoQ -/- mice, there was a significant increase in CPP to methamphetamine (1 mg/kg) but not to a range of d-amphetamine doses (0.5, 1, 2 and 4 mg/kg). Taken together, these data suggest that Cdh13 and AdipoQ regulate sensitivity to psychomotor stimulants and palatable rewards without producing major changes in other behaviors. In humans, these two genes may regulate sensitivity to natural and drug rewards, thus influencing susceptibility to the conditioned drug effects and relapse.

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.

Myopericytoma: a unifying term for a spectrum of tumours that show overlapping features with myofibroma. A review of 14 cases.

BACKGROUND: Myopericytoma (MPC) is a recently proposed term to describe a group of tumours that originate from perivascular myoid cells and show a range of histological growth patterns. Only a small number of series describing MPC have been reported. MPC is frequently misdiagnosed as a sarcoma. AIMS: To document the clinical and histopathological findings of a series of MPCs, to describe the range of growth patterns and morphological spectrum, and to compare MPC with myofibroma (MF). PATIENTS/METHODS: Fourteen patients with features of MPC and/or MF were identified from the archival files of the department of anatomical pathology, Royal Prince Alfred Hospital, Sydney, Australia. RESULTS: There were six female and eight male patients. The mean and median patient ages were 37 and 35.5 years, respectively. The tumours were located in the skin, subcutis, or superficial soft tissues of the distal extremities (13 patients) or the head and neck region (one patient), and showed a spectrum of morphological appearances. They were divided into two groups based upon the predominant growth pattern corresponding to MPC (seven cases) and MF (seven cases). The feature most suggestive of MPC was the presence of a concentric perivascular arrangement of plump spindle shaped cells. The presence of a zonation/biphasic appearance was most characteristic of MF. CONCLUSIONS: MPC exhibits a spectrum of growth patterns that overlap with MF. Tumours can be designated as MPC or MF depending on the predominant growth pattern.

Neuronal overexpression of Glo1 or amygdalar microinjection of methylglyoxal is sufficient to regulate anxiety-like behavior in mice.

GLO1 (Glyoxalase1) is a ubiquitous cellular enzyme that detoxifies methylglyoxal (MG), which is a byproduct of glycolysis. Previously, we showed that ubiquitous overexpression of Glo1 reduced concentrations of MG and increased anxiety-like behavior, whereas systemic injection of MG reduced anxiety-like behavior. We further showed that MG is a competitive partial agonist at GABA-A receptors. Based on those data we hypothesized that modulation of GABAergic signaling by MG underlies Glo1 and MG's effects on anxiety-like behavior. As previous studies used ubiquitous overexpression, we sought to determine whether neuronal Glo1 overexpression was sufficient to increase anxiety-like behavior. We generated ROSA26 knock-in mice with a floxed-stop codon upstream from human Glo1 (FLOXGlo1KI) and bred them with mice expressing CRE recombinase under the direction of the Synapsin 1 promoter (Syn-CRE) to limit overexpression of Glo1 specifically to neurons. Furthermore, since previous administration of MG had been systemic, we sought to determine if direct microinjection of MG into the basolateral amygdala (BLA) was sufficient to reduce anxiety-like behavior. Thus, we performed bilateral microinjections of saline, MG (12µM or 24µM), or the positive control midazolam (4mM) directly into the BLA. FLOXGlo1KIxSyn-CRE mice showed significantly increased anxiety-like behavior compared to their FLOXGLO1xWT littermates. In addition, bilateral microinjection of MG and midazolam significantly decreased anxiety-like behavior compared to saline treated mice. These studies suggest that anatomically specific manipulations of Glo1 and MG are sufficient to induce changes in anxiety-like behavior.

Integration of genome-wide association and extant brain expression QTL identifies candidate genes influencing prepulse inhibition in inbred F1 mice.

Genetic association mapping in structured populations of model organisms can offer a fruitful complement to human genetic studies by generating new biological hypotheses about complex traits. Here we investigated prepulse inhibition (PPI), a measure of sensorimotor gating that is disrupted in a number of psychiatric disorders. To identify genes that influence PPI, we constructed a panel of half-sibs by crossing 30 females from common inbred mouse strains with inbred C57BL/6J males to create male and female F1 offspring. We used publicly available single nucleotide polymorphism (SNP) genotype data from these inbred strains to perform a genome-wide association scan using a dense panel of over 150,000 SNPs in a combined sample of 604 mice representing 30 distinct F1 genotypes. We identified two independent PPI-associated loci on Chromosomes 2 and 7, each of which explained 12-14% of the variance in PPI. Searches of available databases did not identify any plausible causative coding polymorphisms within these loci. However, previously collected expression quantitative trait locus (eQTL) data from hippocampus and striatum indicated that the SNPs on Chromosomes 2 and 7 that showed the strongest association with PPI were also strongly associated with expression of several transcripts, some of which have been implicated in human psychiatric disorders. This integrative approach successfully identified a focused set of genes which can be prioritized for follow-up studies. More broadly, our results show that F1 crosses among common inbred strains can be used in combination with other informatics and expression datasets to identify candidate genes for complex behavioral traits.

Systems genetic and pharmacological analysis identifies candidate genes underlying mechanosensation in the von Frey test.

Mechanical sensitivity is commonly affected in chronic pain and other neurological disorders. To discover mechanisms of individual differences in punctate mechanosensation, we performed quantitative trait locus (QTL) mapping of the response to von Frey monofilament stimulation in BXD recombinant inbred (BXD) mice. Significant loci were detected on mouse chromosome (Chr) 5 and 15, indicating the location of underlying polymorphisms that cause heritable variation in von Frey response. Convergent evidence from public gene expression data implicates candidate genes within the loci: von Frey thresholds were strongly correlated with baseline expression of Cacna2d1, Ift27 and Csnk1e in multiple brain regions of BXD strains. Systemic gabapentin and PF-670462, which target the protein products of Cacna2d1 and Csnk1e, respectively, significantly increased von Frey thresholds in a genotype-dependent manner in progenitors and BXD strains. Real-time polymerase chain reaction confirmed differential expression of Cacna2d1 and Csnk1e in multiple brain regions in progenitors and showed differential expression of Cacna2d1 and Csnk1e in the dorsal root ganglia of the progenitors and BXD strains grouped by QTL genotype. Thus, linkage mapping, transcript covariance and pharmacological testing suggest that genetic variation affecting Cacna2d1 and Csnk1e may contribute to individual differences in von Frey filament response. This study implicates Cacna2d1 and Ift27 in basal mechanosensation in line with their previously suspected role in mechanical hypersensitivity. Csnk1e is implicated for von Frey response for the first time. Further investigation is warranted to identify the specific polymorphisms involved and assess the relevance of these findings to clinical conditions of disturbed mechanosensation.

Behavioral sensitization to ethanol is modulated by environmental conditions, but is not associated with cross-sensitization to allopregnanolone or pentobarbital in DBA/2J mice.

RATIONALE: The ability of ethanol to facilitate GABA(A) receptor-mediated transmission may result in GABA(A) receptor alterations during repeated ethanol administration, and lead to dynamic behavioral changes, including sensitization to the locomotor stimulant effect of ethanol. Since alterations in GABA(A) receptors are likely to alter sensitivity to GABAergic drugs such as 3alpha-hydroxy-5alpha-pregnan-20-one (allopregnanolone) and pentobarbital, we determined whether enhanced sensitivity to ethanol was associated with enhanced sensitivity (cross-sensitization) to these drugs. Two procedures that produced differences in the magnitude of expression of ethanol-induced locomotor sensitization were used. METHODS: After habituation to testing procedures for 2 days, female DBA/2J mice were injected with ethanol or saline for 12 days. On the following day, locomotion was recorded after a challenge injection of ethanol (2 g/kg), allopregnanolone (10 or 17 mg/kg), or pentobarbital (10 or 20 mg/kg). Due to evidence that exposure to the test chambers influenced sensitization, in some experiments, mice were exposed to the test apparatus on the day prior to challenge. RESULTS: Exposure to the test apparatus prior to drug challenge attenuated the expression of ethanol sensitization, compared with mice without this pre-exposure. Cross-sensitization was not observed to either allopregnanolone or pentobarbital under any condition; however, some groups of repeated ethanol-treated mice displayed tolerance to the initial stimulant effects of allopregnanolone and pentobarbital. CONCLUSIONS: These studies indicate that behavioral sensitization to ethanol is not associated with cross-sensitization to pentobarbital or allopregnanolone, and that the expression of ethanol sensitization is influenced by the relative novelty of the test chamber. In addition, these results do not support a mechanism in which alterations in the neurosteroid or barbiturate modulatory sites of the GABA(A) receptor are responsible for the expression of sensitization to the locomotor stimulant effects of ethanol.

Activation of the extracellular signal regulated kinase (ERK) pathway in human melanoma.

BACKGROUND: Several studies suggest that melanoma may be resistant to treatment because of resistance to apoptosis and that this may be the result of activation of the extracellular signal regulated kinase (ERK1/2) pathway. AIMS: To test this hypothesis by examining the expression of ERK1/2 and its activated form in histological sections of melanoma and its relation to known prognostic features of the disease. MATERIALS/METHODS: Immunohistochemistry with antibodies to ERK1/2 and phosphorylated ERK (p-ERK) was performed on formalin fixed sections from 42 primary melanomas, 38 metastases, and 20 naevi. Fourteen of the primary melanomas were in the radial and 28 in the vertical growth phase. RESULTS: ERK1/2 was widely expressed (100%) in all the (pigmented) lesions studied. p-ERK1/2 expression was much lower in compound (32.4%) and dysplastic (54.5%) naevi than in primary melanoma (nodular 78.8%, superficial spreading 67%) and subcutaneous metastases (76.3%). p-ERK expression was much lower in lymph node metastases (48.5%), suggesting that the microenvironment may influence the activation of ERK. There was a (non-significant) trend for p-ERK expression to be higher in thick (>1.0 mm) versus thin (< or =1.0 mm) melanoma (p = 0.23). There was a trend for overall survival to be related to p-ERK expression in patients with melanoma over 1 mm in thickness. CONCLUSIONS: Expression of activated ERK1/2 in melanocytic lesions appears to be related to malignant potential so that activation of ERK1/2 may be important in melanoma progression. These results provide important histological support for the proposal that inhibition of this signalling pathway may be useful in treatment of melanoma.

Bioinformatic analysis of autism positional candidate genes using biological databases and computational gene network prediction.

Common genetic disorders are believed to arise from the combined effects of multiple inherited genetic variants acting in concert with environmental factors, such that any given DNA sequence variant may have only a marginal effect on disease outcome. As a consequence, the correlation between disease status and any given DNA marker allele in a genomewide linkage study tends to be relatively weak and the implicated regions typically encompass hundreds of positional candidate genes. Therefore, new strategies are needed to parse relatively large sets of 'positional' candidate genes in search of actual disease-related gene variants. Here we use biological databases to identify 383 positional candidate genes predicted by genomewide genetic linkage analysis of a large set of families, each with two or more members diagnosed with autism, or autism spectrum disorder (ASD). Next, we seek to identify a subset of biologically meaningful, high priority candidates. The strategy is to select autism candidate genes based on prior genetic evidence from the allelic association literature to query the known transcripts within the 1-LOD (logarithm of the odds) support interval for each region. We use recently developed bioinformatic programs that automatically search the biological literature to predict pathways of interacting genes (PATHWAYASSIST and GENEWAYS). To identify gene regulatory networks, we search for coexpression between candidate genes and positional candidates. The studies are intended both to inform studies of autism, and to illustrate and explore the increasing potential of bioinformatic approaches as a compliment to linkage analysis.

Evidence for differences in cultured left ventricular fibroblast populations isolated from spontaneously hypertensive and Wistar-Kyoto rats.

OBJECTIVE: To compare fibroblast populations derived from spontaneously hypertensive rats (SHRLJ) and normotensive Wistar-Kyoto rats (WKYLJ) for angiotensin II receptor binding, gene expression of the AT1 receptor and angiotensinogen, hormone responsiveness and phenotypic changes. METHODS: Fibroblasts were isolated by either collagenase B or collagenase P and grown to confluency in the presence of 10% fetal bovine serum. Angiotensin II receptor binding was assessed under both serum and serum-free conditions. Hormonal treatment of cells was conducted in a serum-free background. The concentrations of AT1 receptor and angiotensinogen messenger RNA (mRNA) were determined by liquid hybridization. Phenotypic changes in fibroblast populations were analysed by visualization of lipid-containing vacuoles (oil red O stain) or of alpha-smooth muscle actin-containing fibres (immunostain). RESULTS: SHRLJ collagenase-B cells grew more slowly and had nearly twofold fewer angiotensin II receptors than WKYLJ cells as measured by both radioligand binding and AT1 mRNA content (SHRLJ 1.34 +/- 0.05 versus WKYLJ 5.94 +/- 0.41 pg mRNA per microgram total RNA) but contained significantly more angiotensinogen mRNA (SHRLJ 147 +/- 12 versus WKYLJ 98 +/- 8 fg mRNA per microgram total RNA). Collagenase-P cells from the two strains exhibited similar binding and growth properties. Collagenase-B fibroblasts also exhibited greater responses to exogenous steroids, including a greater shift towards an adipocyte phenotype, than collagenase-P cells. Exogenous angiotensin II promoted transformation towards a myofibroblast cell type, especially in collagenase-P SHRLJ cells. CONCLUSION: Our results indicate that subsets of fibroblasts that differ in growth rate, angiotensin II receptor binding, AT1 and angiotensinogen mRNA levels, structure and steroid responsiveness may be isolated from the left ventricle. The potential importance of these altered phenotypes to cardiac remodelling and hypertrophy warrants further examination.