Generalized Membrane Defect Research Articles

Non-retinal Abnormalities Associated with Progressive Retinal Atrophy (PRA) in the Miniature Poodle

Osmotic fragility was examined in red blood cells taken from miniature poodles with progressive retinal atrophy (PRA). In this study, the median osmotic fragility for erythrocytes (concentration of NaCl required for 50% haemolysis) was found to be significantly lower for clinically affected, compared to unaffected animals. These differences were maintained regardless of changes in erythrocyte incubation pH (7.0, 7.5). There was no difference in the slope parameter of erythrocyte osmotic fragility profiles for both sets of animals, indicating a similar variability of erythrocyte age. In groups of animals maintained with the same diet and fasted for 12 hr, and in groups of animals with no dietary constraints, total plasma cholesterol was significantly lower for affected, compared to unaffected animals. Red blood cell indices were also assessed; mean corpuscular volume (MCV), mean corpuscular haemoglobin (MCH), red blood cell count (RBC) and blood haemoglobin concentration (Hb) were normal, while erythrocyte packed cell volume (PCV) was significantly reduced for affected, compared to unaffected. Data from this study suggest that the abnormal erythrocyte osmotic fragility of affected animals is unlikely to be a consequence of a higher erythrocyte surface area/volume ratio, higher plasma cholesterol concentration, or greater proportion of new to old erythrocytes, and that a general membrane defect might be associated with prcdin the miniature poodle.

Calcium homeostasis in fibroblasts from patients with amyotrophic lateral sclerosis

Amyotrophic lateral sclerosis (ALS) is a chronic neurodegenerative disorder of the motor system in the CNS characterized by motor neuron death in the spinal cord, brain stem and cortex. Readily available tissues such as fibroblasts from ALS patients can serve as simple model systems to study the molecular mechanisms leading to degenerative disorders. We have used Fura-2 fluorescence microscopy and single-cell imaging to study the spatiotemporal dynamics of intracellular free calcium ([Ca 2+] i) in primary cultures of fibroblasts from skin biopsies from ALS and normal subjects. Increases in [Ca 2+] i were induced by stimulation with bradykinin (100 nM); neurotensin (50 nM); N-formyl-Met-Leu-Phe (chemotactic peptide) (1 μM); [Arg 8]-vasopressin (1 μM) and histamine (10 μM). The levels of [Ca 2+] i in 80–120 individual cells per agonist were monitored for 15 min. No significant differences were found in the resting levels of [Ca 2+] i in control (102 ± 4 nM) and ALS (98 ± 6 nM) fibroblasts and in the maximal [Ca 2+] i levels after stimulation with N-formyl-Met-Leu-Phe, [Arg 8]-vasopressin, and histamine. Significantly lower [Ca 2+] i transients were found in fibroblasts from ALS donors compared to controls when stimulated with neurotensin ( p < 0.002) and bradykinin ( p < 0.005). The percentage of individual cells reacting to a given agonist (40–100%) was similar in both groups. The molecular basis of the impaired calcium homeostasis in fibroblasts from ALS patients is not known, but a generalized membrane defect can be excluded since the [Ca 2+] i responses are defective only when bradykinin or neurotensin are used as agonists.

Analysis of lipid composition of sarcoplasmic reticulum membranes from normal and malignant hyperthermic pig skeletal muscle.

In search of a general membrane defect hypothesis for malignant hyperthermia syndrome, we analysed the lipid profiles of heavy sarcoplasmic reticulum membranes isolated from normal and malignant hyperthermia longissimus dorsi pig muscle. Malignant hyperthermia susceptibility was assessed by halothane challenge of pigs. Sarcoplasmic reticulum membranes from malignant hyperthermia susceptible pigs differed significantly from control ones in the cholesterol content and phosphatidylethanolamine/phosphatidylcholine ratio; both were higher in former membranes. These latter lipid modifications were in agreement with the significant increase of their bulk lipid viscosity, as evidenced by an increase of diphenyl hexatriene fluorescence anisotropy. The increased level of phosphatidylethanolamine associated with the decreased content of phosphatidylcholine in malignant hyperthermic membranes was shown to be a potential consequence of depressed activities of both phospholipid N-methyltransferase I and II activities. Finally, the distribution of fatty acids in these particular phospholipids showed no change in phosphatidylcholine molecules, whereas the percentage of arachidonate and stearate in the phosphatidylethanolamine species were respectively higher and lower in malignant hyperthermic membranes. These differences in major phospholipids content and the enrichment of a metabolically important fatty acyl chains in malignant hyperthermia sarcoplasmic reticulum membranes strongly suggest that the lipid metabolism may contribute to the molecular mechanism of malignant hyperthermia syndrome.

Chloride transport in control and cystic fibrosis human skin fibroblast membrane vesicles

Plasma membrane vesicles were isolated from either cystic fibrosis (CF) or non-CF cultured fibroblasts derived from skin biopsies of either foetus, child or adolescent human donors. The total membrane yield was essentially identical for either CF or control membranes. By using a rapid filtration technique, 36Cl uptake by these vesicles was quantitated in the absence and presence of alkali-metal ion-, electrical- and/or pH gradients. In the absence of a pH gradient (pH out = pH in = 7.5), Cl uptake took place downhill in both cases. Either cis K +, cis Na + or an equimolar mixture of cis Na + plus K + caused Cl uptake activation. In the presence of an alkaline-inside pH gradient (pH out/pH in = 5.5/7.5), Cl uptake exhibited an apparent overshoot independently of the presence or absence of any metal-ion gradient. The observed potassium-, sodium- and proton-dependent Cl influx rates were all unaffected by voltage clamping, indicating the existence in these vesicles of electroneutral symport systems of the type Cl −/H +, Cl −/K + and/or Cl −/Na +; but not 2 Cl −/Na +/K +. In the presence of an inward-directed K + gradient, valinomycin further increased Cl uptake, bot in the presence and in the absence of a pH gradient, indicating the presence of a rheogenic Cl uniport. In absolute quantitative terms, the two different modes (rheogenic and electroneutral) of Cl transport evinced in these vesicles were about 45% lower in CF than in control skin fibroblasts. However, qualitatively, there was no difference between normal and CF cells. The evidence obtained indicates that the CF defect, which is expressed in fibroblast plasma membranes, does not affect specifically either the rheogenic or the electroneutral Cl transport systems. Rather, the CF cells appear to give a smaller yield to closed, functional vesicles, reflected by a significantly smaller apparent intravesicular volume. Because it also affects the transport of d-glucose and l-alanine, this anomaly could be the consequence of a generalized membrane defect characterizing CF fibroblasts.

Abnormalities in the regulation of blood platelet free cytosolic calcium in malignant hyperthermia I. Human platelets

The effect of halothane on the regulation of blood platelet free cytosolic calcium was investigated in Quin-2-loaded cells from patients susceptible to Malignant Hyperthermia (MH) and healthy controls, respectively. The resting level of free cytosolic calcium was slightly, but statistically significantly, enhanced in platelets from patients (90 ± 10 nM vs 110 ± 35 nM). Halothane induced a dose-dependent, rapid Ca 2+ release from intracellular stores both in normal and in MH derived cells, but the resulting increase in cytosolic calcium was significantly higher in the latter (2 mM halothane: [Ca 2+] i = 117 ± 12 nM vs 218 ± 117 nM; 4 mM halothane: 225 ± 35 nM vs. 417 ± 201 nM). Whereas in platelets from healthy donors a complete reversibility of the halothane effect could be observed within 30–45 min, the cytosolic Ca 2+ transients in platelets from patients were different from those in normals either in a higher initial peak or in a diminished decline velocity or in both. The basal Ca 2+ permeability of the platelet plasma membrane was very low. Generally, halothane caused a dose-dependent increase in Ca 2+ permeability. However, the influx of external calcium was significantly higher in platelets from patients than in controls (2 mM halothane: Δ[Ca 2+] i = 69 ± 12 nM vs 135 ± 63 nM; 4 mM halothane: 127 ± 33 nM vs. 258 ± 111 nM). Combining the results, the suggestion can be made that susceptibility to MH is characterized by a generalized membrane defect. A fast halothane-induced intracellular Ca 2+ release and an increased Ca 2+ permeability of the plasma membrane seem to be of particular importance for adverse cellular reactions on exposure to the anaesthetic.

Erythrocyte membrane fluidity in malignant hyperthermia

Malignant hyperthermia (MH) is a rare but serious complication of general anesthesia that potentially carries a high mortality and morbidity. It is associated with excessive release of calcium into skeletal muscle following exposure to certain drugs, including the volatile general anesthetics. Since these are recognized membrane fluidizing agents it has been speculated that this condition might represent a generalized defect in membrane physical properties either at rest or inducible by fluidizing agents. If this hypothesis were found to be correct, malignant hyperthermia might conveniently be detected by examining membrane physical properties of easily accessible cells rather than the cumbersome method of muscle biopsy currently employed. To test this hypothesis we identified patients proven to be susceptible to MH by muscle biopsy and a cohort of patients not susceptible to MH as defined by negative muscle biopsy testing. Erythrocytes were isolated from both groups and membrane physical properties examined using conventional, widely available, steady-state fluorescence polarization techniques. Erythrocyte membranes were evaluated with multiple probes both in the basal condition and following fluidization with either increasing temperature or two concentrations of a fluidizing alcohol. We report, contrary to previous publications, that no discernable differences were detectable between MH-positive or negative patients. Thus, we find no evidence for a generalized membrane defect in MH and conclude that the determination of erythrocyte membrane physical properties, by these techniques, are of no use in the preoperative screening for this disorder.

Adrenocorticotropin Hyperresponsiveness in Myotonic Dystrophy Following Oral Fenfluramine Administration

Abstract The plasma immunoreactive adrenocorticotropin and cortisol responses to oral fenfluramine hydrochloride (1.5mg/kg body wt) or placebo were examined in 11 patients with myotonic dystrophy, 4 controls with facioscapulohumeral dystrophy, a similarly debilitating muscle wasting disease, and 14 normal controls in single-blind studies performed in mid-afternoon. Mean areas under the adrenocorticotropin response versus time curve were significantly greater in myotonics (2573 + 429 pmol.min/L) than in facioscapulohumeral dystrophy controls (696 + 279 pmol.min/L, P<0.02) and normals (560+/-61 pmol.min/L, P<0.0001). Corresponding cortisol responses were significantly greater in myotonics (35757 + 3949 nmol.min/L) than in normals (21828+/-1669 nmol.min/L, P < 0.001), but not significantly greater than those in facioscapulohumeral dystrophy controls (22830 + 6140 nmol.min/L, P = 0.055). No stressful side-effects which could affect hormone responses, and no significant changes in blood pressure or heart rate were noted. Fenfluramine activates central serotonergic and/or noradrenergic pathways initiating secretion of corticotropin-releasing hormone and possibly arginine vasopressin. We postulate that these fenfluramine-activated pathways are hyperstimulated in myotonics, leading to adrenocorticotropin and cortisol hypersecretion. This may be a manifestation of a general cell membrane defect in myotonic dystrophy. We found a lack of correlation of age (and severity of disease) with adrenocorticotropin response in myotonics, and therefore, the hyperresponse may serve as a useful marker for the disease before development of other overt signs.

Abnormal fluidity state in membranes of malignant hyperthermia pig skeletal muscle

The fluidity state was analyzed on sarcoplasmic reticulum membranes and phospholipid vesicles prepared from normal and malignant hyperthermia susceptible pig muscle. Electron spin resonance studies were performed to determine the fluidity state at the region near the polar headgroups and in the central core of the bilayer using 5-nitroxide (5-NS) and 16-nitroxide stearic acid (16-NS), respectively. With the 5-NS label, no differences were found between normal and malignant hyperthermia sarcoplasmic reticulum (MH SR) membranes whereas with the 16-NS label, a significant increase of the activation energy was shown with MH membranes. Lower values of fluorescence anisotropy observed with DPH-labeled MH membranes as compared with normal ones, confirmed the higher abnormal fluidity state of these membranes. The fluidizing effect of halothane, a triggering agent of malignant hyperthermia syndrome, was also studied in these membranes. We show that a relatively low concentration of the drug destabilized not only the diseased sarcoplasmic reticulum membranes but also the vesicles made of total phospholipids extracted from MH skeletal muscle. Together, these findings strongly suggest that an overall increase in membrane fluidity may be implied in the MH disease, improving the general membrane defect hypothesis for this syndrome.

An electron paramagnetic resonance study of skeletal muscle membrane fluidity in malignant hyperthermia

Skeletal muscle sarcolemma (SL), transverse tubule (TT) and heavy sarcoplasmic reticulum (HSR) membranes were isolated from malignant hyperthermia susceptible (MHS) and normal pigs, and the rotational dynamics of lipid hydrocarbon chain motion was examined by electron paramagnetic resonance (EPR) spectroscopy. The stearic acid spin probe 16-SASL was incorporated into MHS and normal membranes and both the order parameter ( S) and effective correlation time ( τ r) of probe motion were calculated from spectra recorded over the temperature range of 2 to 40°C. At any given temperature, TT membranes exhibited significantly greater values for both the S and τ r of probe motion than did SL, which exhibited significantly greater values than did HSR membranes. The order of decreasing S and τ r values for 16-SASL mobility correlated with the decreasing cholesterol content of these membranes (TT>SL>HSR), however there was no difference in the S or τ r values for a given membrane fraction isolated from both MHS and normal muscle. Arrhenius plots of 16-SASL mobility in SL, TT and HSR were linear from 2 to 40°C, indicating no abrupt thermotropic change in the lipid hydrocarbon phase of any of the membrane types studied. Apparent activation energies ( E a), calculated from the Arrhenius plots, were similar for MHS and normal membranes derived from a given cellular location. However, the E a of probe motion for TT membranes (2.3 ± 0.1 and 2.4 ± 0.1 kcal/mol/degree for MHS and normal, respectively) was significantly less than for SL (3.4 ± 0.4 and 2.9 ± 0.1 kcal/mol/degree for MHS and normal, respectively) which, in turn, was significantly less than the E a for HSR (3.7 ± 0.1 and 3.7 ± 0.1 kcal/mol/degree for MHS and normal, respectively). Since 16-SASL motion was similar in MHS and normal membranes, we conclude that there is no evidence for a generalized membrane defect affecting lipid mobility in these MHS muscle membranes.

Plasma Membrane and its Abnormalities in Hypertension

The plasma membrane is composed of proteins embedded in a discontinuous fashion in a lipid bilayer. These proteins maintain the integrity of the membrane and play fundamental roles as ion transport channels and as receptors for agents that regulate cell function. The membrane is therefore an important regulator of vascular smooth muscle contraction. The plasma membrane in the hypertensive animal exhibits abnormal permeability for monovalent ions and defective calcium handling. This is reflected in fewer calcium-binding sites and, as a result, in deficient membrane stabilization. These defects have been identified in several cell types, including lymphocytes, red blood cells, adipocytes, and vascular smooth muscle cells. Evidence presented in the current review suggests that hypertension is associated with a generalized membrane defect. Abnormalities in ion transport in vascular smooth muscle cells are the most relevant to the pathogenesis of hypertension since they could be directly responsible for the rise in blood pressure. We hypothesize that the impaired stabilizing effect of calcium in vascular smooth muscle cells of hypertensive subjects renders the membrane more excitable and that this in turn leads to increased vascular reactivity and higher peripheral resistance. Peripheral vascular reactivity usually is increased in hypertension, suggesting increased responsiveness of the smooth muscle cells. Possible abnormalities of the several components of the contractile process of these cells have been investigated for the role they might play in this altered response. Abnormalities in the plasma membrane have been most clearly defined and are emphasized in this review.

Free cytoplasmic Ca++ at rest and after cholinergic stimulus is increased in cultured muscle cells from Duchenne muscular dystrophy patients.

We used a fluorescent dye, quin 2, to measure intracellular free calcium ([Ca++]i) in cultured skeletal muscle cells and skin fibroblasts from five Duchenne muscular dystrophy (DMD) patients and from five controls. We observed an enhanced [Ca++]i level, at rest and after acetylcholine (ACh) stimulation, in DMD muscle cells, but we did not detect any difference between DMD and normal skin fibroblasts. The abnormally higher [Ca++]i transient induced by ACh suggests that it plays a critical role in muscle degeneration. The skin fibroblast results suggests that there is no generalized membrane defect.

Temperature-dependent abnormalities of the erythrocyte membrane in porcine malignant hyperthermia.

The temperature dependence of ATPase activities and stearic acid spin label motion in red blood cells of normal and MH-susceptible pigs have been examined. Arrhenius plots of red blood cell ghost Ca-ATPase and calmodulin-stimulable Ca-ATPase activities were identical for both normal and MH erythrocyte ghosts. Arrhenius plots of Mg-ATPase activity exhibited a break (defined as a change in slope) at 24 degrees C in both MH and normal erythrocyte ghosts. However, below 24 degrees C the apparent activation energy for this activity was less in MH than normal ghosts. To determine whether breaks in ATPase Arrhenius plots could be correlated with changes in the physical state of the red blood cell membrane, the spin label 16-doxyl-stearate was introduced into the bilayer of both erythrocyte ghosts and red blood cells. With both ghosts and intact cells, at each temperature examined, the mobility of the probe in the lipid bilayer, as measured by electron paramagnetic resonance, was greater in normal than in MH membranes. While there were no breaks in Arrhenius plots for probe motion in the erythrocyte ghosts, the apparent activation energy for probe motion was significantly greater in normal than in MH ghost membranes. While there was no break in the Arrhenius plot of probe motion in normal intact red blood cell membranes, there were breaks in the Arrhenius plot of probe motion at both 24 and 33 degrees C in intact MH red blood cell membranes. Based on the altered temperature dependence of Mg-ATPase activity and spin probe motion in membranes derived from MH red blood cells, we conclude that there may be a generalized membrane defect in MH pigs which is reflected in the red blood cell as an altered membrane composition or organization.

Membrane fluidity of nonmuscle cells in Duchenne muscular dystrophy: effect on lymphocyte membranes of incubation in patient and control sera.

The membrane fluidity of intact fibroblasts, erythrocyte ghosts, and intact lymphocytes from Duchenne muscular dystrophy (DMD) patients and controls was measured by steady state fluorescence polarization. The fluorescent probes used were diphenylhexatriene (DPH), trimethylammonium-DPH, and a set of n-(9-anthroyloxy) fatty acids. Fluorescence anisotropies in DMD fibroblasts and DMD erythrocyte ghosts were normal. In DMD lymphocytes (n = 10) fluorescence anisotropy of DPH was decreased versus controls (0.212 +/- 0.028 versus 0.231 +/- 0.012, p less than 0.05). Linear regression analysis of creatine kinase activity in sera and DPH fluorescence anisotropy in lymphocytes from DMD patients showed a negative correlation (r = -0.93, p less than 0.001). DMD lymphocytes and control lymphocytes were incubated for 4 h in sera from DMD patients and from controls. When incubated in DMD sera, DPH fluorescence anisotropy of DMD lymphocytes decreased from 0.211 +/- 0.018 to 0.180 +/- 0.028, and fluorescence anisotropy of control lymphocytes decreased from 0.239 +/- 0.012 to 0.179 +/- 0.025 reaching the same level as did DMD lymphocytes. When incubated in control sera, DPH fluorescence anisotropy of DMD lymphocytes increased to 0.224 +/- 0.012, and fluorescence anisotropy of control lymphocytes decreased to 0.218 +/- 0.017. The fluorescence anisotropy changes after incubation in DMD versus control sera were different (p less than 0.05 for DMD lymphocytes and p less than 0.005 for control cells). Our findings do not support the hypothesis of a general membrane defect but suggest a toxic serum factor in DMD which attacks lymphocyte membranes and possibly muscle membranes at the same time.

Alterations in lipid membrane fluidity and the physical state of cell-surface sialic acid in zinc-deficient rat erythrocyte ghosts

Erythrocyte ghosts, prepared from the blood of rats fed zinc-deficient diets, were evaluated for membrane fluidity and surface sialic acid properties using spin-labeled probes and electron spin resonance (ESR) spectroscopy. These physical parameters of the erythrocyte ghosts from the zinc-deficient group were compared to those for erythrocyte ghosts obtained from ad libitum and pair fed controls consuming zinc-adequate diets. As the animals became progressively zinc deficient, the erythrocyte ghost membranes became more fluid than those from the control groups. In addition, the apparent rotational correlation time of Tempamine spin probes on surface sialic acid residues was smaller for the zinc deficient group, indicative of an increased rotational mobility of the spin label. These results suggest that zinc deficiency can have pronounced effects on the physical state of membrane bilayer lipids and cell surface carbohydrates and supports the view that many of the pathological signs of zinc deficiency are due to a general membrane defect.

Some biophysical properties of the erythrocyte membrane in Duchenne muscular dystrophy

Inasmuch as prior authors have suggested a ‘generalized membrane defect’ in patients with Duchenne muscular dystrophy, this study was undertaken to examine some biophysical properties of the erythrocytes in affected individuals. Using the micropipette technique we investigated the rheological properties of erythrocytes from 11 Duchenne muscular dystrophy patients (boys 3–14 years old) and 5 carriers (mothers 20–42 years old). The mean elastic modulus and membrane viscosity of the patient's erythrocytes were not significantly different from those of a group of 8 unaffected age-matched boys. The carrier group did not differ significantly from the adult control group in any of the rheological parameters. A significant increase in the number of stomatocytes was observed in the patient group, and a similar trend (not statistically significant) was observed in the carrier group. The membrane lipid fluidity of erythrocytes from patients, carriers, and normal controls was assessed with three different probes by fluorescence depolarization, and no significant differences were found. Our results indicate that there is a progressive elevation of the membrane elastic modulus with increasing age of the patients: the younger patients have a lower elastic modulus than their age-matched controls, while the older patients have a higher elastic modulus than their age-matched controls.

Freeze-fracture analysis of intramembrane particles of erythrocytes from normal and dystrophic hamsters.

The intramembrane particles on plasma membranes of erythrocytes of Syrian hamsters afflicted with hereditary muscular dystrophy were compared to those from normal controls by freeze-fracture analysis. The reduced number of particles in both fracture faces of dystrophic erythrocyte plasmalemmae, as compared to that of control red cells, was found to be highly significant (P = .001). Results of this study therefore, support the concept of a generalized membrane defect (abnormality) in muscular dystrophy.

Estrone treatment dissociates primary versus secondary consequences of "diabetes" (db) gene expression in mice.

Feeding 0.001% estrone in a diet to C57BL/KsJ mice homozygous for the recessive obesity gene "diabetes" (db) permitted dissociation of the primary consequences of obesity gene expression from the secondary consequences of diabetes effected through interaction between the db gene and other diabetogenic genes in the inbred background. Estrone-treated db/db mice were similar to untreated mutants in exhibiting hyperphagia and marked obesity. However, estrone-treated mutants did not develop the hyperinsulinemia, hyperglycemia, and islet atrophy characteristic of untreated db/db mice. Thus, expression of the primary defect could be studied in the absence of the myriad secondary sequelae elicited by chronic hyperinsulinemia and hyperglycemia. Reduced numbers of hepatocyte plasma membrane insulin receptors (50% of normal) persisted in the estrone-treated mice in the absence of hyperinsulinemia, indicating that this deficiency was a consequence of the primary genetic defect and not merely a downregulation phenomenon secondary to hyperinsulinemia. Comparison of insulin secretion from comparably sized +/+ islets versus islets from estrone-treated db/db mice showed no intrinsic defects in beta-cell sensitivity to glucose. In conclusion, db-induced obesity can be dissociated from hyperinsulinemia, hyperglycemia, beta-cell dysfunction, and hyperphagia but is associated with a generalized membrane defect reflected in part by the persistent deficiency of plasma membrane insulin receptors.

Spin label study of red blood cell membranes in Huntington's disease.

Huntington's disease (HD) is an inherited degenerative disorder of the central nervous system. Various studies of peripheral tissues from HD patients have led to the assumption that this disease could be the expression of a generalized membrane defect. We have been unable to reproduce fundamental results on which this theory is based, namely differences between electron spin resonance spectra for erythrocytes from diseased and healthy persons. Contradictory results have been published for other methods as well so that it is necessary to rethink the membrane defect theory.

Abnormal erythrocyte and lymphocyte chemiluminescence in chronic granulomatous disease: Evidence for a generalized membrane defect

Concanavalin A (soluble)- and/or opsonized zymosan (particulate)-stimulated luminol-dependent chemiluminescence was demonstrated in normal human lymphocytes, erythrocytes, and in the membrane (ghost) fractions of polymorphonuclear neutrophils and erythrocytes. By contrast, seven patients with chronic granulomatous disease (five boys with the X-linked type and two female variants) had diminished luminescent responses to con A and/or opsonized zymosan in their lymphocytes, erythrocytes, erythrocyte membranes, as well as neutrophil membranes, suggesting generalization of the deficiency in the neutrophil oxidase system to other cells.

31P-NMR studies of normal and dystrophic chicken muscle.

Phosphorus-31 nuclear magnetic resonance (31P-NMR) analysis was performed on normal (line 412) and dystrophic (line 413) superficial pectoralis muscles excised from chickens between 15 and 116 days after hatching. An apparent alteration in dystrophic muscle energy metabolism was abolished by pretreatment with curare and was attributed to muscle hyperexcitability. Time-dependent 31P-NMR studies demonstrated no apparent difference in the overall tissue adenosine triphosphatase (ATPase) activity of dystrophic as compared to normal muscle. After 55 days of age, a resonance signal attributed to serine ethanolamine phosphodiester (SEPDE) was observed only in dystrophic muscle. Adding the paramagnetic cation Mn2+ to the buffer surrounding the muscle resulted in an approximate 80% decrement in the dystrophic SEPDE signal without apparent alteration of the other phosphatic signals in either the normal or dystrophic muscle. This would argue against any generalized membrane defect in dystrophic chicken muscle and suggest that SEPDE is in a compartment accessible to Mn2+.