Association study of serine racemase gene with methamphetamine psychosis.

Experimental studies have demonstrated that not only dopaminergic signaling but also glutamatergic/NMDA receptor signaling play indispensable roles in the development of methamphetamine psychosis. Our recent genetic studies provided evidence that genetic variants of glutamate-related genes such as DTNBP1, GLYT1, and G72, which are involved in glutamate release and regulation of co-agonists for NMDA receptors, conferred susceptibility to methamphetamine psychosis. Serine racemase converts l-serine to d-serine, which is an endogenous co-agonist for NMDA receptors. Three single nucleotide polymorphisms (SNPs) in the promoter region of the serine racemase gene (SRR), rs224770, rs3760229, and rs408067, were proven to affect the transcription activity of SRR. Therefore, we examined these SNPs in 225 patients with methamphetamine psychosis and 291 age- and sex-matched controls. There was no significant association between methamphetamine psychosis and any SNP examined or between the disorder and haplotypes comprising the three SNPs. However, rs408067 was significantly associated with the prognosis for methamphetamine psychosis and multi-substance abuse status. The patients with C-positive genotypes (CC or CG) of rs408067 showed better prognosis of psychosis after therapy and less abuse of multiple substances than the patients with GG genotypes. Because the C allele of rs408067 reduces the expression of SRR, a lower d-serine level or reduced NMDA receptor activation may affect the prognosis of methamphetamine psychosis and multiple substance abuse. Our sample size is, however, not large enough to eliminate the possibility of a type I error, our findings must be confirmed by replicate studies with larger samples.

Association study between casein kinase 1 epsilon gene and methamphetamine dependence.

Casein kinase 1 epsilon (CKIepsilon) is a component of the DARPP-32 in second-messenger pathway. CKIepsilon phosphorylates and activates DARPP-32, a key molecule in various complex signaling pathways, including dopamine and glutamine signaling, which have both been demonstrated to be main pathways in substance dependence. A recent clinical study showed that rs135745, a noncoding single nucleotide polymorphism of the 3'-untranslated region of the CSNK1E gene, was associated with the intensity of the subjective response to an oral amphetamine dose in normal volunteers. Differences in sensitivity to the drug should affect development of dependence to it. Hence, we genotyped rs135745 of the CSNK1E (MIM 600863) gene in 215 patients with methamphetamine dependence and 274 age- and gender-matched normal controls. No significant differences in genotype and allele frequencies were observed between the patients with methamphetamine dependence and controls. There was also no significant association between rs135745 and the clinical characteristics of methamphetamine dependence and co-morbid methamphetamine psychosis (e.g., age of first consumption, latency of psychosis, prognosis of psychosis after therapy, spontaneous relapse of psychotic symptoms, and poly-substance abuse status). The present findings suggest that having a genetic variant of the CSNK1E gene did not affect susceptibility to methamphetamine dependence or psychosis, at least in a Japanese population.

Biased sinusoidal field gel electrophoresis for the separation of large DNA.

In agarose gel electrophoresis, in a steady, continuous field, it is well known that the mobility mu, versus size M relation for linear DNAs (L-DNAs) can be divided into three regimes: Ogston regime I for small DNAs, where M dependence of mu, is weak; entangled but unstretched regime II for intermediate-size L-DNAs (of M < 20 kbp), where mu, sigma M-1 so that efficient fractionation is possible; and entangled and stretched regime III for large L-DNAs, where M dependence of mu s is again weak. Although mu s and the regimen boundaries can be altered by adjusting the gel concentration Cgel and/or the field strength E, the features of the M dependence of mu s are essentially unchanged. As to the effect of DNA topology on mu s, we found that in dilute gels (Cgel < 1.0 wt%) coiled, circular DNAs (C-DNAs) of 2-7 kbp size migrate faster than L-DNAs of comparable size, while in concentrated gels (Cgel > 1.5 wt%) C-DNAs migrate much slower than L-DNAs. To facilitate separation of large DNAs in the regime III range, we proposed biased sinusoidal field gel electrophoresis (BSFGE), which utilizes a sinusoidal field of strength Es and frequency f superposed on a steady bias field of strength Eb.(ABSTRACT TRUNCATED AT 250 WORDS)

Cell adhesion onto block copolymer Langmuir-Blodgett films.

The attachment of cells onto the surfaces of various block copolymers fabricated either as well-defined, ordered Langmuir-Blodgett (LB) films, or solvent cast microphase-separated structures was studied. In general, more platelets adhered onto the multilayered LB surface than onto microphase-separated cast surfaces. Scanning electron micrographs of adhered platelets showed extensive morphological changes associated with the LB surface as compared to cast film surfaces. The morphology of adhered hepatocytes was similar for both LB films and cast surfaces. It may be assumed that the surface of a block copolymer LB film does not orient into microdomains, as in the solvent cast surfaces, and only one polymer domain interacts at the interface.

The determination of molecular weights of Streptomyces subtilisin inhibitor and the complex of Streptomyces subtilisin inhibitor and subtilisin BPN' by sedimentation equilibrium.

The molecular weight of Streptomyces subtilisin inhibitor (SSI), a protein proteinase inhibitor, and that of the complex of SSI and subtilisin BPN' [EC 3.4.21.14] were determined by a sedimentation equilibrium method in 25 mM phosphate buffer, at pH 7.0, ionic strength 0.1 M (NaCl), 25.0 degrees C. The molecular weight of SSI was found to be 23,000 over a wide concentration range, 0.01-10 mg/ml, the range used for inhibitory, spectrophotometric, and kinetic measurements. Based on the amino acid sequence, the molecular weight of SSI has been calculated to be 11,500 (Ikenaka, T., et al. (1974) J. Biochem. 76, 1191-1209); therefore, the molecular weight of 23,000 obtained above suggests that SSI is in a dimeric form under usual conditions in the concentration range of 5 X 10(-7)-5 X 10(-4) M. The molecular weight of the subtilisin BPN'-SSI complex was determined to be 78,000 in the concentration range of 0.03-5.0 mg/ml by sedimentation equilibrium of the crystallized preparation and by that of a mixture of subtilisin BPN' and SSI treated as a multicomponent-polydisperse system. The molecular weight obtained here, combined with the results of binding stoichiometry (Inouye, K., et al. (1977) J. Biochem. 82, 961-967) that showed that one mol of SSI (molecular weight, 11,500) and one mol of the enzyme (molecular weight, 27,500) are tightly bound (Kd less than 1 nM), demonstrate that one mol of dimeric SSI binds two mol of the enzyme to form a stable complex, E2I2.