Measuring electrostatic potentials adjacent to membranes.
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Biomedical subjects
Publications and source records attributed to A McLaughlin.
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We investigated the binding of physiologically and pharmacologically relevant ions to the phosphoinositides by making 31P NMR, electrophoretic mobility, surface potential, and calcium activity measurements. We studied the binding of protons to phosphatidylinositol 4,5-bisphosphate (PIP2) by measuring the effect of pH on the chemical shifts of the 31P NMR signals from the two monoester phosphate groups of PIP2. We studied the binding of potassium, calcium, magnesium, spermine, and gentamicin ions to the phosphoinositides by measuring the effect of these cations on the electrophoretic mobility of multilamellar vesicles formed from mixtures of phosphatidylcholine (PC) and either phosphatidylinositol, phosphatidylinositol 4-phosphate, or PIP2; the adsorption of these cations depends on the surface potential of the membrane and can be described qualitatively by combining the Gouy-Chapman theory with Langmuir adsorption isotherms. Monovalent anionic phospholipids, such as phosphatidylserine and phosphatidylinositol, produce a negative electrostatic potential at the cytoplasmic surface of plasma membranes of erythrocytes, platelets, and other cells. When the electrostatic potential at the surface of a PC/PIP2 bilayer membrane is -30 mV and the aqueous phase contains 0.1 M KCl at pH 7.0, PIP2 binds about one hydrogen and one potassium ion and has a net charge of about -3. Our mobility, surface potential, and electrode measurements suggest that a negligible fraction of the PIP2 molecules in a cell bind calcium ions, but a significant fraction may bind magnesium and spermine ions.
Human gastrocnemius and slow twitch muscles contain phosphodiesters that may be detected in vivo by phosphorus nuclear magnetic resonance (NMR). This work represents a study of 354 spectra obtained from healthy subjects of various ages and from patients with peripheral vascular disease. The analysis of the data indicate a correlation between the concentration of phosphodiesters and age. By comparing the data obtained with healthy subjects and patients it is concluded that the increase in phosphodiesters is not due to disease, but to ageing itself. The significance of this increase is discussed.
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We measured the electrophoretic mobility of multilamellar phospholipid vesicles, the 31P NMR spectra of both sonicated and multilamellar vesicles, and the conductance of planar bilayer membranes to study the binding of spermine and gentamicin to membranes. Spermine and gentamicin do not bind significantly to the zwitterionic lipid phosphatidylcholine. We measured the concentrations of gentamicin and spermine that reverse the charge on vesicles formed from a mixture of phosphatidylcholine and either phosphatidylserine or phosphatidylinositol. From these measurements, we determined that the intrinsic association constants of the cations with these negative lipids are all about 10 M-1. This value is orders of magnitude lower than the apparent binding constants reported in the literature by other groups because the negative electrostatic surface potential of the membranes and the resultant accumulation of these cations in the aqueous diffuse double layer adjacent to the membranes have not been explicitly considered in previous studies. Our main conclusion is that the Gouy-Chapman-Stern theory of the aqueous diffuse double layer can describe surprisingly well the interaction of gentamicin and spermine with bilayer membranes formed in a 0.1 M NaCl solution if the negative phospholipids constitute less than 50% of the membrane. Thus, the theory should be useful for describing the interactions of these cations with the bilayer component of biological membranes, which typically contain less than 50% negative lipids. For example, our results support the suggestion of Sastrasinh et al. [Sastrasinh, M., Krauss, T. C., Weinberg, J. M., & Humes, H. D. (1982) J. Pharmacol. Exp. Ther. 222, 350-358] that phosphatidylinositol is the major binding site for gentamicin in renal brush border membranes.
Serial exercise thallium-201 myocardial perfusion scanning (exercise and 4-hour redistribution) was compared to rest and exercise electrocardiography (ECG) for the detection of coronary artery disease in 125 patients with known or suspected coronary artery disease. All patients underwent coronary arteriography and 108 were found to have significant coronary artery lesions. The serial exercise thallium scan was significantly more sensitive than rest and exercise ECG in detecting coronary artery disease (94% v. 83% P less than 0.01). The sensitivity of a reversible thallium perfusion scan abnormality and a positive exercise ECG for detecting exercise induced myocardial ischaemia in coronary artery disease was similar (69% v. 63%). The exercise thallium scan complemented the exercise ECG, and the sensitivity of the combined test was significantly greater than the exercise ECG alone (84% v. 63% P less than 0.001). The specificity for coronary artery disease of the exercise ECG was 65% and that of the exercise thallium-201 myocardial perfusion scan was 82% (P = NS). Thallium-201 myocardial perfusion scanning complements the rest and exercise ECG in the non-invasive detection of coronary artery disease.
Electrophoretic mobility and 31P NMR measurements were combined to test whether the combination of the Henry, Boltzmann and Grahame equations is capable of describing the adsorption of divalent cations of phosphatidylcholine membranes. Cobalt was chosen for this study because, of all the common divalent cations, its effects on the 31P NMR spectrum of phosphatidylcholine membranes are easiest to interpret. Both the 31P NMR data on the adsorption of cobalt and the zeta potential data calculated from the electrophoretic mobility in the presence of cobalt are well described by the combination of these three equations. Electrophoretic mobility measurements were also performed with a number of other divalent cations and the zeta potentials were, in all cases, well described by the combination of these three equations. The binding deduced from such measurements decreases in the sequence: Mn2+, Mg2+, Ca2+, Co2+, Ni2+, Sr2+, Ba2+. If we assume that a lipid molecule occupies an area of 60 A2 and that there is a 1 : 1 stoichiometry for the binding of the divalent ions to phosphatidylcholine, the dissociation constants are, respectively: 0.3, 1.0, 1.0, 1.2, 1.2, 2.8, 3.6 M.
Thallium--201 myocardial perfusion scanning has been evaluated in Australia in patients with coronary artery disease. Myocardial scans reliably detect both acute myocardial infarction and the transient myocardial ischaemia of angina pectoris. The non-invasive nature, ease of study, and the ability to scan patients with conventional cameras makes thallium--201 an attractive additional diagnostic agent for patients with suspected coronary artery disease. Although thallium--201 reliably indicates perfusion defects in the myocardium, its diagnostic use at the moment should be reserved to clarify such diagnostic problems in patients with coronary artery disease which cannot be satisfactorily explained by conventional investigation.
Technetium 99m Pyrophosphate imaging before and after open heart surgery was performed in 38 patients to estimate the incidence of peri-operative infarction. Positive images were present pre-operatively in 11 of 30 patients with coronary artery disease. In three patients the images changed from negative to positive and in two this was thought to be due to infarction produced at operation. The high incidence of positive pre-operative images emphasises that many patients with evolving myocardial infarction, unstable angina or severe heart failure are now operated upon without delay. Because many patients have positive images before surgery pre-operative and post-operative images must be compared to assess myocardial damage due to surgery.
Twenty-nine patients with suspected or actual myocardial infarction have had myocardial "hot spot" scans with technietium 99m pyrophosphate, using a mobile nuclear camera in a coronary care unit. Nine patients with transmural infarcts had positive scans. Nine out of 12 patients thought to have had endocardial infarction had positive scans. Two patients who had had intramuscular injections presented with chest pain and abnormally high serum enzyme levels, but had negative scans. Myocardial radionuclide scanning with a mobile nuclear camera is a rapid, easy, repeatable, non-invasive method of helping to confirm or exclude the diagnosis of myocardial infarction and, used together with other available tests, is a valuable additional diagnostic aid.
In a series of eight patients with the Sturge-Weber syndrome, the brain scan was shown to be the most accurate, non-invasive, diagnostic test, being abnormal in all eight patients. In four cases examined by cerebral arteriography, this study was also conspicuously abnormal. Some of the neurological disturbances occurring in these patients are more likely to be due to transient ischaemic attacks and cerebral infarctions than to epilepsy.
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