Semantic priming and verbal learning in current opiate users, ex-users and non-user controls.

OBJECTIVE: Despite a growing interest in memory functions of chronic drug users, investigation of semantic and episodic memory in opiate users is limited, and findings of studies have been inconsistent. The present study aimed to assess semantic memory and episodic memory for both drug-related and neutral stimuli in current and ex-users of opiates. METHODS: Using an independent group design, we assessed semantic priming and verbal learning in 16 current opiate users on a methadone maintenance programme, 16 ex-opiate users in rehabilitation programmes and 16 healthy controls. The groups were matched on verbal IQ, age and employment status. RESULTS: We found that current and ex-users showed intact automatic and controlled semantic priming. Ex-users who had been abstinent for an average of 19 months showed a verbal learning impairment compared with controls. Both current and ex-users were impaired in recalling semantically unrelated words but unimpaired in recalling semantically related words. CONCLUSION: The findings suggest a relative lack of spontaneous use of mnemonic strategies and imply that highly structured information would help opiate-using clients in treatment.

Attentional bias, inhibitory control and acute stress in current and former opiate addicts.

BACKGROUND: Drug dependence is associated with both attentional biases to drug-related cues and inhibitory control deficits. Although acute stress is known to increase craving, it is not known whether this effect is mediated via changes in attentional bias and inhibitory control. OBJECTIVES: To examine the effect of a mild stressor on inhibitory control, attentional bias and craving in current opiate users (methadone maintained), ex-users (currently abstinent) and non-users (healthy controls). METHOD: Forty-eight participants (16 in each group) were exposed to both stress and non-stress conditions, after which inhibitory control and attentional bias was assessed using a Go-No-go and dot probe task respectively. Subjective ratings of stress levels and drug craving were repeatedly monitored. RESULTS: Current opiate users had significantly higher cravings ratings than both other groups at all times, and their craving tended to increase following the stress task. Current users had a greater attentional bias towards drug-related stimuli than the ex-users. Interestingly, ex-users showed a bias away from drug-related stimuli in the stress condition and this correlated positively with their length of abstinence. On the Go/No-go task, all groups had fewer false alarms in the stress condition. CONCLUSIONS: These results indicate that successful treatment is associated with a bias away from drugs, and that this bias may be protective against the effects of stress.

Genotyping beta-globin gene mutations on copolymer-coated glass slides with the ligation detection reaction.

BACKGROUND: Methods are needed to analyze small amounts of samples for variation in disease-causing genes. One means is to couple the sensitivity and multiplexing capability of the ligation detection reaction (LDR) with the use of simple glass slides specifically functionalized with a novel polymer coating to enhance sensitivity. METHODS: We developed an array-based genotyping assay based on glass slides coated with copolymer (N,N-dimethylacrylamide, N,N-acryloyloxysuccinimide, and 3-(trimethoxysilyl)propyl methacrylate). The assay consists of an LDR with genomic DNA followed by a universal PCR (U-PCR) of genomic DNA-templated LDR product. The LDR occurs in the presence of 3 primers for each sequence variant under investigation: 2 distinguishing primers (allele specific and perfectly complementary to wild-type and mutant alleles) and 1 common locus-specific primer. The 2 allele-specific primers have different capture sequences for binding different complementary probes on a tag array. The LDR product templated from genomic DNA is made fluorescent during the U-PCR via incorporation of a Cy5-labeled universal primer into all LDR products; detection occurs on the coated glass slides. RESULTS: The assay was designed to detect 7 prevalent mutations in the beta-globin gene (HBB, hemoglobin, beta) in a multiplex format, and signals for the different alleles are detected by their fluorescence. The assay was applied to 40 genomic DNA samples from both control individuals and patients with known beta-thalassemia mutations. Results show good correspondence between the patients' genotypes as assessed by DNA sequence analysis and those generated from the LDR assays. CONCLUSIONS: The developed technology allows accurate identification of sequence variants in a simple, cost-effective way and offers good flexibility for scaling to other applications with different numbers of single-nucleotide polymorphisms or mutations to be detected.

De novo deletion removes a conserved motif in the C-terminus of ABCA4 and results in cone-rod dystrophy.

BACKGROUND: Mutations in the retina-specific ABC transporter (ABCA4) gene are associated with different types of macular degeneration, including Stargardt disease, cone-rod dystrophy, Fundus flavimaculatus, Retinitis pigmentosa and probably age-related macular degeneration. METHODS: Screening for mutations in the ABCA4 gene was performed using denaturing high-performance liquid chromatography and direct sequencing. RESULTS: We describe the identification of a new de novo 44-bp deletion in an Italian patient affected by cone-rod dystrophy. The mutation, located in intron 48 of the ABCA4 gene, is predicted to cause exon 49 skipping, resulting in loss of the C-terminus of the ABCA4 protein. Interestingly, exon 49 also codes for a highly conserved VFVNFA motif, which has been demonstrated to be essential for the activity of ABCA1, another gene of the ABC transporter family. The presence of CT repeats at the breakpoints might have facilitated the generation of the deletion through a slippage mispairing mechanism. CONCLUSIONS: The new 6730-16del44 deletion is the first de novo mutation associated with cone-rod dystrophy and may contribute to a better understanding of the role of ABCA4 mutations in macular dystrophies.

Denaturing HPLC profiling of the ABCA4 gene for reliable detection of allelic variations.

BACKGROUND: Mutations in the retina-specific ABC transporter (ABCA4) gene have been associated with several forms of macular degenerations. Because the high complexity of the molecular genotype makes scanning of the ABCA4 gene cumbersome, we describe here the first use of denaturing HPLC (DHPLC) to screen for ABCA4 mutations. METHODS: Temperature conditions were designed for all 50 exons based on effective separation of 83 samples carrying 86 sequence variations and 19 mutagenized controls. For validation, samples from 23 previously characterized Stargardt patients were subjected to DHPLC profiling. Subsequently, samples from a cohort of 30 patients affected by various forms of macular degeneration were subjected to DHPLC scanning under the same conditions. RESULTS: DHPLC profiling not only identified all 132 sequence alterations previously detected by double-gradient denaturing gradient gel electrophoresis but also identified 5 sequence alterations that this approach had missed. Moreover, DHPLC scanning of an additional panel of 30 previously untested patients led to the identification of 26 different mutations and 29 polymorphisms, accounting for 203 sequence variations on 29 of the 30 patients screened. In total, the DHPLC approach allowed us to identify 16 mutations that had never been reported before. CONCLUSIONS: These results provide strong support for the use of DHPLC for molecular characterization of the ABCA4 gene.