Islets in most species respond to increased glucose with biphasic insulin secretion, marked by a sharp first-phase peak and a slowly rising second phase. Mouse islets in vitro, however, lack a robust second phase. To date, this observation has not been extended in vivo. We thus compared insulin secretion from conscious mice with isolated mouse islets in vitro. The arterial plasma insulin response to a hyperglycemic clamp was measured in conscious mice 1 wk after surgical implantation of carotid artery and jugular vein catheters. Mice were transfused using clamps with blood from a donor mouse to maintain blood volume, allowing frequent arterial sampling. When plasma glucose in vivo was raised from ϳ5 to ϳ13 mM, insulin rose to a first-phase peak of 403 Ϯ 73% above basal secretion (n ϭ 5), followed by a rising second phase of mean 289 Ϯ 41%. In contrast, perifused mouse islets (ϳ75 islets/trial) responded with a similar first phase of 508 Ϯ 94% (n ϭ 4) but a smaller and virtually flat second phase of 169 Ϯ 9% (n ϭ 4, P Ͻ 0.05). Furthermore, the slope of the second-phase response differed significantly from zero in mice (2.63 Ϯ 0.39%/min, P Ͻ 0.01), in contrast to perifused islets (0.18 Ϯ 0.14%/min, P Ͼ 0.30). Mice also displayed pulsatile patterns in insulin concentration (period: 4.2 Ϯ 0.4 min, n ϭ 8). Conscious mice thus responded to increased glucose with biphasic and pulsatile insulin secretion, as in other species. The robust second phase observed in vivo suggests that the processes needed to generate second-phase insulin secretion may be abrogated by islet isolation.insulin; secretion; first phase; second phase; biphasic; mouse; in vivo; pulses; pulsatile INSULIN SECRETION OCCURS IN A BIPHASIC MANNER in response to rapid increases in glucose concentration (2,8,20,45). Firstphase secretion consists of a sharp peak that occurs within 2-5 min of a glucose challenge (29). It is generally accepted that the first-phase peak results from the glucose stimulus increasing -cell ATP/ADP to close ATP-dependent K ϩ (K ATP ) channels, and in turn, depolarizing the -cell membrane potential. This results in the opening of voltage-gated calcium channels, promoting the influx of calcium and secretion of an immediately releasable pool of granules (15,37,46). Secondphase secretion consists of a sustained elevation of insulin secretion, which rises over a period of several minutes (2,8,20,45) and is typically accompanied by insulin pulses occurring at 5-to 15-min intervals (34).This description of biphasic insulin secretion is typical of nearly all species, whether measured from in vivo plasma concentrations (6, 7), perfused pancreas (8, 12), or perifused islets in vitro (21). A notable exception is the isolated mouse islet, which displays a small and relatively static second phase after a fairly typical first-phase response. This type of flat second phase has been observed using a variety of mouse preparations, including the perfused pancreas and cultured islets, and is seen in several different mouse strains (4,13,23,26,42,5...