Complex repetitive arrangements of gene sequence in the candidate region of the spinal muscular atrophy gene in 5q13.

Childhood-onset proximal spinal muscular atrophy (SMA) is a heritable neurological disorder, which has been mapped by genetic linkage analysis to chromosome 5q13, in the interval between markers D5S435 and D5S557. Here, we present gene sequences that have been isolated from this interval, several of which show sequence homologies to exons of beta-glucuronidase. These gene sequences are repeated several times across the candidate region and are also present on chromosome 5p. The arrangement of these repetitive gene motifs is polymorphic between individuals. The high degree of variability observed may have some influence on the expression of the genes in the region. Since SMA is not inherited as a classical autosomal recessive disease, novel genomic rearrangements arising from aberrant recombination events between the complex repeats may be associated with the phenotype observed.     

SR141716A, a potent and selective antagonist of the brain cannabinoid receptor

SR141716A is the first selective and orally active antagonist of the brain cannabinoid receptor. This compound displays nanomolar affinity for the central cannabinoid receptor but is not active on the peripheral cannabinoid receptor. In vitro, SR141716A antagonises the inhibitory effects of cannabinoid receptor agonists on both mouse vas deferens contractions and adenylyl cyclase activity in rat brain membranes. After intraperitoneal or oral administration SR141716A antagonises classical pharmacological and behavioural effects of cannabinoid receptor agonists. This compound should prove to be a powerful tool for investigating the in vivo functions of the anandamide/cannabinoid system.     

In Memoriam

The Patient Self-Determination Act was implemented in December 1991. Before and after its implementation, we used a structured interview of 302 randomly selected patients to determine their awareness, understanding, and use of advance directives. Implementation of the Act did not have a major effect on these. Although more than 90% of patients were aware of the living will, only about a third selected the correct definition or the correct circumstances in which it applied, and less than 20% of patients had completed one. About a third of patients were aware of a Durable Power of Attorney for Health Care and chose the correct definition, and about half identified the correct circumstances in which it applies; less than 10% had completed such a document. Surprisingly, patients who said they had completed advance directives did not demonstrate better understanding of these documents. Our results indicate that many patients, including some who have completed advance directives, do not fully understand them. It may be unwise to regard these documents as carefully considered, compelling statements of patients'' preferences. Appropriate responses to our findings include increased public education, revising state statutes to bring them into congruence with public perception, and expanding the dialogue between physicians and patients.     

Aluminium polyphosphate complexes in the mycorrhizal basidiomyceteLaccaria bicolor: A27Al-nuclear magnetic resonance study

The techniques of ²⁷Al- and ³¹P-nuclear magnetic resonance (NMR) spectroscopy were used to investigate the interactions of aluminium with intracellular ligands within the mycelium of the ectomycorrhizal basidiomycete Laccaria bicolor (Maire) Orton (S238). The vegetative mycelium was grown on medium containing 0.5 mM AlCl₃ for 0.5 to 3 d. The ²⁷Al-NMR spectra showed that aluminium was rapidly taken up and accumulated into polyphosphate complexes in the vacuole. Comparison with Al-polyphosphate complexes obtained in vitro on model systems indicated that Al forms at least three mixed-solvation complexes with Pi and polyphosphates, that there is more than one complex present under any set of conditions, and that the equilibrium between these complexes shifts dramatically with Al concentration in the medium. The high phosphate concentrations in the growth medium favoured the accumulation of the Al-polyphosphate complexes. When mycelium containing Al-polyphosphate complexes was transferred to Al-free nutrient solution for 9 d, the Alpolyphosphate complexes were not remobilized. The sequestration of Al in the polyphosphate complexes could therefore make a significant contribution to the protection of mycorrhizal plants against aluminium toxicity.Abbreviations NMR nuclear magnetic resonance - PolyP polyphosphate(s) - PP₁ terminal phosphate of PolyP - PP₃ middle phosphate of PolyP We thank Prof. Daniel Canet (Laboratoire de Méthodologie RMN, University of Nancy I, Vandceuvre-lès-Nancy, France) for his constant encouragement and Christine Delaruelle for skilled technical assistance in growing the fungal cultures. This work was supported by a research grant from the Commission of the European Communities (STEP-CT90-0059, Role of Ectomycorrhiza in Stress Tolerance of Forest Trees) to F.M. and a travel grant from the Institut National de la Recherche Agronomique to I.K.; R.C. is a recipient of a Postdoctoral Fellowship from the Natural Sciences and Engineering Research Council of Canada.     

Refining the genetic map for the region flanking the X-linked hypophosphataemic rickets locus (Xp22.1–22.2)

An analysis of five of the most common cystic fibrosis (CF) mutations worldwide (F-508, R-553X, G-551D, N-1303K and G-542X) was performed in 36 Chilean patients. Polymerase chain reaction (PCR) amplification of the DNA followed by allele specific restriction enzyme analysis was used for detection. The overall frequencies of the mutations in the chromosomes analyzed were 29.2% for F-508 and 4.2% for R-553X (n=72). The G-542X, G-551D and N-1303 K mutations were absent in the Chilean sample. Our data suggest however that F-508 is not the most common CF mutation in Chilean patients. F-508 and R-553X account for only 33.4% of the alleles; 66.6% of them do not respond to the probes used and still remain uncharacterized.     

Oxidants act as chemorepellents in Paramecium by stimulating an electrogenic plasma membrane reductase activity

Paramecium is a valuable eukaryotic model system for studying chemosensory transduction, adaptation and cellular sensory integration. While millimolar amounts of many attractants hyperpolarize and cause faster forward swimming, oxidants are repellents that depolarize and cause backward swimming at micromolar concentrations. The non-permeant oxidants cytochrome c, nitro blue tetrazolium and ferricyanide are repellents with half maximal concentrations of 0.4 M, 2.2 M and 100 M respectively. In vivo reductase activities follow the same order of potencies. The concentration dependence of the cytochrome c reductase activity is well correlated with cytochrome c-induced depolarizations. This suggests that plasma membrane reduction of external cytochrome c is electrogenic, causing membrane depolarization and chemorepulsion. The reductase activity also appears to be voltage dependent. Depolarization by either K+, Na+, Ca+ or Mg+ correlates with inhibition of both in vivo reductase activities and cytochrome c-induced membrane potential changes. These responses were also seen in deciliated cells, showing that the body plasma membrane is sufficient for the response. Both chloroquine and diphenyleneiodonium inhibited reductase activities but only at unusually high concentrations. This activity showed no pH dependence in the physiological range. We propose that a plasma membrane bound NADPH-dependent reductase controls oxidant-induced depolarizations and consequent chemorepulsion.Abbreviations bmv Body plasma membrane vesicles - BPS Bathophenanthroline disulfonate - cAMP Cyclic adenosine monophosphate - cmv Ciliary membrane vesicles - cyt c Cytochrome c - DPI Diphenyleneiodonium - EC ₅₀ Concentration for 50% effectiveness - FeCN Ferricyanide [Fe(CN)₆–3] - FeEDTA Ethylenediaminetetracetic acid (ferric-sodium salt) - GTP Guanosine 5-triphosphate - KCN Potassium cyanide - mM Millimolar - MOPS 3-(N-morpholino) propanesulfonic acid - mV Millivolts - NADH Nicotinamide adenine dinucleotide (reduced form) - NADPH Nicotinamide adenine dinucleotide phosphate (reduced form) - NBT Nitro blue tetrazolium - nm Nanometer - pCMB p-Chloromercuribenzoate - PMA Phorbol 12-myristate 13-acetate - s.d. Standard deviation - SOD Superoxide dismutase - Tris Tris(hydroxymethyl)aminomethane - M Micromolar