. The Biological bulletin. Biology; Zoology; Marine biology. 90 M. E. KRIEBEL ET Figure 1. Dissociated electrocytes from Raja erinacea after low-temperature incubation with coUagenase. Note the range of sizes of these whole, unfixed electrocytes as seen in the dissecting microscope. Electrocyte E, is about one-third the size of E2. NE = nerve ending cluster. Bar = mm. Enzyme assays Lactate dehydrogenase (LDH) was measured by fol- lowing the oxidation of NADH spectrophotometrically (Johnson, 1960) in a final volume of 1 ml. Acetylcholin- esterase (AChE) was measured using a final incub
. The Biological bulletin. Biology; Zoology; Marine biology. 90 M. E. KRIEBEL ET Figure 1. Dissociated electrocytes from Raja erinacea after low-temperature incubation with coUagenase. Note the range of sizes of these whole, unfixed electrocytes as seen in the dissecting microscope. Electrocyte E, is about one-third the size of E2. NE = nerve ending cluster. Bar = mm. Enzyme assays Lactate dehydrogenase (LDH) was measured by fol- lowing the oxidation of NADH spectrophotometrically (Johnson, 1960) in a final volume of 1 ml. Acetylcholin- esterase (AChE) was measured using a final incubation volume of 1 ml (Ellman 1961). Choline acetyltrans- ferase (ChAT) was used as a marker for nerve terminal cytoplasm and was measured by means of a radiometric assay described by Fonnum (1975), with the following modifications. The final incubation volume of 10-100 ^1 contained NaCl (300); EDTA (20); choHne chloride (10); acetyl CoA (); eserine sulfate (); sodium phosphate buffer, pH (50); Triton X-100; and ^H- or "Re- labeled acetyl CoA (500 nCi/ml). Incubations were at 26°C for time periods up to 2 h, and three time points were used for each determination. Reactions were ter- minated by transfer of aliquots (5-50 ^1) to mini-scintil- lation vials containing ml of 10 mA/phosphate buffer and ml acetonitrile containing sodium tetraphenyl- boron (10 mg/ml). Labeled ACh was measured by liquid scintillation after addition of ml of water-immiscible scintillation fluid, gentle shaking, and phase separation. Labeled acetyl CoA remained in the lower (aqueous) layer and therefore made no contribution to the count rate. This procedure gave the same results as one in which the volumes of reagents after termination were 3 times greater (as originally described by Fonnum, 1975). Both coUa- genase and skate saline were found to inhibit ChAT ac- tivity when added to extracts of whole electric organ. This inhibitory effect was therefore minimized b
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