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Myotonic dystrophy (DM) is one of the most prevalent muscular diseases in adults. The molecular basis of this autosomal disorder has been identified as the expansion of a CTG repeat in the 3' untranslated region of a gene encoding a protein kinase (DMPK). The pathophysiology of the disease and the role of DMPK are still obscure. It has been previously demonstrated that DMPK is localized at neuromuscular junctions, myotendinous junctions, and terminal cisternae of the sarcoplasmic reticulum (SR), in the skeletal muscle, and at intercalated discs in the cardiac muscle. We report here new findings about specific localization of DMPK in the heart. Polyclonal antibodies raised against a peptide sequence of the human DMPK were used to analyze the subcellular distribution of the protein in rat papillary muscles. Confocal laser microscopy revealed a strong although discontinuous reactivity at intercalated discs, together with transverse banding on the sarcoplasm. At higher resolution with immunogold electron microscopy, we observed that DMPK is localized at the cytoplasmic surface of junctional and extended junctional sarcoplasmic reticulum, suggesting that DMPK is involved in the regulation of excitation-contraction coupling. Along the intercalated disc, DMPK was found associated with gap junctions, whereas it was absent in the two other kinds of junctional complexes (fasciae adherentes and desmosomes). Immunogold labeling of gap junction purified fractions showed that DMPK co-localized with connexin 43, the major component of this type of intercellular junctions, suggesting that DMPK plays a regulatory role in the transmission of signals between myocytes.

Dystrophin, the product of the Duchenne muscular dystrophy gene, has been shown to be developmentally regulated in both human muscle and brain tissues. We consequently performed an immunocytochemical study using electron microscopy to localise the protein in the immature human fetal muscle and neurons. Results demonstrated that, even if dystrophin was partially associated to the plasma membrane in both tissues, some product was also linked to the neurofilaments network in neurons and to microfilaments in muscle. An intense staining was also found in satellite cells.

The present experiments were designed to test acetylcholine (ACh) release from cholinergic nerve endings of the chicken iris during aging. Either a one-step K+ stimulated release scheme or a more strenuous two-step depletion-reloading-release scheme were used. First, we found that independent of the scheme of release, as demonstrated by a lower peak release, slower time to peak and a lower rate of increase, the 3-year-old iris released less 3H-ACh than the 4-month-old iris. In order to analyze this effect of aging, we determined five basic parameters of ACh metabolism, including total acetylcholine (ACh) and choline (Ch) levels, 3H-ACh, 3H-Ch and 3H-phosphorylcholine (PhCh) formed from exogenous 3H-Ch, for each experimental type and for the two ages. There appear to be major differences in the handling of both the total Ch and newly labelled (3H-Ch) pools between the two ages. In addition, the homeostatic control system for Ch levels in the older tissue appears to be less efficient and incapable of handling strenuous release tests. This defect of the older tissue to preserve its Ch pool may, in part, be responsible for the drastic decrease in ACh levels seen in the older iris. Together with the demonstrated reduction in neurotransmitter level and in uptake of Ch, the reduction in release demonstrated here forms a characteristic triade of age-related defects which may have important implications for our understanding of aging processes in cholinergic terminals.

The immunological reactivity of isolated sarcoplasmic reticulum from rabbit fast muscle was tested with antibody to the Ca2+-pump protein which is the predominant component of these membranes. Microplate enzyme-linked immunoassay (ELISA) gave highly reproducible results under the conventional conditions used for checkerboard titration of soluble antigens and antibody. Parallel electron microscope observation of the incubated SR vesicles, negatively stained with ammonium molybdate, shows that the immunologically reactive form of the Ca2+-pump protein is still present in membrane-bound form.