Identification of the milk fat globule membrane proteins. II. Isolation of major proteins from electrophoretic gels and comparison of their amino acid compositions

The fat globule membranes of milk are derived from the apical plasma membrane of the mammary secretory cells. The nature of the membrane proteins, as isolated from cows' milk, has been studied by the use of discontinuous and continuous SDS-gel electrophoresis. Six methods of preparation of milk fat globule membrane suggested by various authors were tested; gel electrophoresis showed that five major bands were present, independent of the method of preparation. The apparent molecular masses of these proteins as determined on SDS-gels (15% T) were 167, 142, 64, 49 and 46 kDa, respectively. The 167 kDa band stained only with periodic acid-Schiff reagent, while the 142 kDa band stained only with Coomassie blue; the last three bands stained with both. Delipidated membranes were extracted stepwise with water, 0.02 M NaCl and 0.6 M NaCl. The 64 kDa band appears to be nearly insoluble, while the bands of 142, 49 and 46 kDa are fractionated by this procedure. The resolution of all of these proteins by electrophoresis was superior to that achieved by molecular sieve chromatography, and so electrophoretic extraction was used to isolate the major proteins. Dansyl chloride derived proteins were used as markers. Amino acid compositions of the recovered proteins were obtained and are compared.     

Changes in insulin response to glucose after exercise training in partially pancreatectomized rats

The effects of exercise training on glucose-stimulated insulin secretion (GSIS) were studied in male Sprague-Dawley rats made mildly to severely diabetic by partial pancreatectomy. Exercise trained (10 wk treadmill; T) and untrained (Unt) rats were grouped according to posttraining fed-state hyperglycemia as follows: T less than 200 and Unt less than 200 (glucose concn less than 200 mg/dl), T 200-300 and Unt 200-300 (glucose concn 200-300 mg/dl), and T greater than 300 and Unt greater than 300 (glucose concn greater than 300 mg/dl). After exercise training, hyperglycemic glucose clamps were performed in awake rats by elevation of arterial blood glucose concentration 126 mg/dl above fasting basal levels for 90 min. Exercise training significantly increased muscle citrate synthase activity. Prevailing hyperglycemia was reduced during the 10-wk exercise training period in all T rats with fed-state glucose concentrations less than 300, and only 53% of Unt rats in these groups had reduced glycemia. GSIS was significantly higher in T less than 200 [2.4 +/- 0.7 (SD) ng/ml at 90 min] than in Unt less than 200 (1.5 +/- 0.3). A similar response was found for T 200-300 (1.1 +/- 0.3 ng/dl) vs. Unt 200-300 (0.7 +/- 0.1) but not T greater than 300 (0.36 +/- 0.2) vs Unt greater than 300 (0.44 +/- 0.05). Sham-operated control rats had insulin concentrations of 6.6 +/- 1.6 ng/ml at the 90th min of the clamp. Acute exercise reduced fed-state glycemia in rats with mild-to-moderate (less than 300 mg/dl) diabetes.(ABSTRACT TRUNCATED AT 250 WORDS)     

Kinetics of release of serotonin from isolated secretory granules. I. Amperometric detection of serotonin from electroporated granules.

We developed a method for measuring the efflux of 5-hydroxytryptamine (5-HT, serotonin) from isolated intact granules of the mast cell of the beige mouse. This method combines electroporation of the vesicle membrane with amperometric detection of 5-HT. A single secretory granule is placed between two platinum electrodes (distance approximately 100 microm) and positioned adjacent (<1 microm) to a carbon fiber microelectrode. A short (approximately 30 micros) high-intensity voltage pulse (electric field of approximately 5 kV/cm) is delivered to the electrodes to trigger the mechanical breakdown of the granule membrane, which activates the release of 5-HT. We observed concurrent swelling of the granule matrix with the oxidation of 5-HT at the carbon fiber electrode (overpotential + 650 mV). Similar to the release of secretory products during exocytosis, the oxidation current exhibits a spike-like time course with a noninstantaneous rising phase (time between onset of current and maximum flux, t(max)) with approximately 25% of the molecules released during this period. When the current reaches its maximum, the granule matrix attains its maximum swollen state. We found that the rising phase depends on the initial cross-sectional area of the granule (t(max) approximately 21r2) and reflects the time required for membrane rupture. The average t(1/2)spike of the amperometric spikes was found to be approximately 150 ms, which is 3-7 times faster than the t(1/2) measured during cellular exocytosis.     

Rinvenimenti Floristici Marchigiani

Summary

The author communicates the list of taxa not commonly occurring in the “Marche”, which he has collected in localities not yet mentioned in the bibliography of that area. The list includes also several species more or less widely cultivated in that zone.