Three minutes after the application of “type”:”entrez-nucleotide”,”attrs”:”text”:”A23187″,”term_id”:”833253″,”term_text”:”A23187″A23187, digitonin (100?M) was added to produce cell lysis and so allow the total available K+ to be estimated (Cook & Haylett, 1985). and so allow the total available K+ to be estimated (Cook & Haylett, 1985). The magnitude of the K+ loss initiated by “type”:”entrez-nucleotide”,”attrs”:”text”:”A23187″,”term_id”:”833253″,”term_text”:”A23187″A23187 could then be calculated as the increase in [K]0 3?min after addition of “type”:”entrez-nucleotide”,”attrs”:”text”:”A23187″,”term_id”:”833253″,”term_text”:”A23187″A23187, expressed as a percentage of the total increase after addition of digitonin. This is equivalent to the quantity of K+ released by “type”:”entrez-nucleotide”,”attrs”:”text”:”A23187″,”term_id”:”833253″,”term_text”:”A23187″A23187 as a percentage of total K+ content of the cells. The inhibitory effects of PK(Ca)-blocking drugs were tested by adding a small volume (usually 5?l) of a concentrated stock treatment for the cell suspension for a preincubation period (usually 3?min) before applying “type”:”entrez-nucleotide”,”attrs”:”text”:”A23187″,”term_id”:”833253″,”term_text”:”A23187″A23187 to initiate K+ loss Ca2+-activated K+-channels. The loss of K+ in the presence of the drug was then compared with that in its absence, so that the inhibition caused by the drug could be expressed as a percentage. Much longer preincubation periods (up to 2?h) were explored in some experiments. In these instances, packed red cells (20?l) were added to a glass vial containing 2?ml of the standard low K+ answer containing the drug. The vial was gently shaken in a water bath at 37C for the time required. Its contents were then transferred to the recording chamber prior to the application of A23817. Because the K+ content of the incubation ISA-2011B fluid was constantly monitored, the rate at which IL4 the cells lost K+ when treated with “type”:”entrez-nucleotide”,”attrs”:”text”:”A23187″,”term_id”:”833253″,”term_text”:”A23187″A23187 could also be determined. This was done by expressing the amount (Q) of K+ lost during successive 20?s periods as a fraction of the K+ content (Q) of the cells midway in that period. Dividing this fraction by the time (t, normally 20?s) over which the loss occurred provided an estimate of the rate coefficient (is percentage inhibition, is a rate constant and is time (see also Table 1). The onset of the action of nitrendipine was too rapid to be resolved by present technique and the broken line has been constructed using a value of of 7?min?1, to indicate a lower limit. Though the factors that underlie the slow onset of action of the cetiedil series have not been studied in any detail, the onset was noted to be approximately exponential in time course, with a rate constant that increased with the activity of the compound. Table 1 lists the rate constants for cetiedil, UCL 1269 and UCL 1274 together with the concentrations causing half maximal inhibition (IC50). Because the potency of these substances is strongly correlated ISA-2011B with their lipophilicity (Benton the anion exchanger (Simons, 1984) and to activate the Ca2+-dependent K+ channels by a direct effect not involving Ca2+ (Shields em et al /em ., 1985). In keeping with this, the addition of Pb2+ to rabbit erythrocytes suspended in the standard low K+ answer caused a loss of K+ comparable to, though a little slower than, that seen with “type”:”entrez-nucleotide”,”attrs”:”text”:”A23187″,”term_id”:”833253″,”term_text”:”A23187″A23187. In six such experiments, the mean K+ loss in response to a 5-min ISA-2011B application of Pb2+ at 10?M (a maximal concentration) was 531%, as compared with 58.45% ( em n /em =6) with the standard application of “type”:”entrez-nucleotide”,”attrs”:”text”:”A23187″,”term_id”:”833253″,”term_text”:”A23187″A23187 (2?M, also maximal). As physique 7a shows, the cetiedil congener UCL 1274 was as effective in blocking K+ loss induced by Pb2+ as by “type”:”entrez-nucleotide”,”attrs”:”text”:”A23187″,”term_id”:”833253″,”term_text”:”A23187″A23187. The IC50’s observed in this set of experiments were 5.40.4?M (with Pb2+) and 5.30.4?M (“type”:”entrez-nucleotide”,”attrs”:”text”:”A23187″,”term_id”:”833253″,”term_text”:”A23187″A23187). Open in a separate window Physique 7 (a) Inhibition by UCL 1274 of K+ loss from rabbit erythrocytes exposed to either A23187 (2?M) or Pb2+ (10?M). Each point is the mean of 3C4 observations and.
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