Comparison of amperometric methods for detection of exocytosis from singlepancreatic beta-cells of different species

Two methods for amperometric detection of exocytosis at single pancreatic b eta-cells were compared. In the first, direct detection of insulin was acco mplished using an insulin-sensitive chemically modified electrode. In the s econd, 5-hydroxytryptamine (5-HT) that had been allowed to accumulate withi n the beta-cell secretory vesicles was detected with a bare carbon electrod e, The goal of the comparison was to determine whether 5-HT secretion was a valid marker of insulin secretion in single beta-cells. To aid in this com parison, some experiments involved simultaneous measurement of insulin and 5-HT at cells previously allowed to accumulate 5-HT. Upon application of co mmon insulin secretagogues, current spikes resulting from detection of 5-HT , insulin, or both compounds were obtained indicative of secretion via exoc ytosis. The mean area of current spikes obtained from simultaneous measurem ents equaled the sum of the mean area of insulin and 5-HT measured independ ently. Additionally, analyses of the number of spikes obtained for detectio n of insulin, 5-HT, or both compounds were similar for several common secre tagogues. These data support the hypothesis that accumulated 5-HT is releas ed from insulin containing secretory vesicles, exclusively. In addition, me asurement of insulin and 5-HT from beta-cells of different species was comp ared to determine whether a species dependence exists for the two methods c ompared here. Detection of 5-HT results in a similar number of spikes that are equivalent to insulin in frequency and amplitude in human, porcine, and canine beta-cells; however, in mouse and INS-1 beta-cells, 5-HT is more re adily detected than insulin.