Figure 1.

ChR2 activation creates local [Ca²⁺]_i elevation in pancreatic islets. (A) A model of spatial activation of ChR2 in pancreatic islets. Activation regions within the islet are defined, where ChR2 is activated by 458 nm illumination leading to depolarization and opening of V-gated Ca²⁺ channels. (B) An activation region defined over the whole islet (left, green dashes) generates islet-wide [Ca²⁺]_i influx (left, orange). [Ca²⁺]_i increases rapidly after ChR2 activation, as measured by Rhod-2 fluorescence (right). (C) An activation region defined over a quadrant of the islet (left, green dashes) generates local [Ca²⁺]_i elevation, which extends outside of the activation region (left, orange). More distant areas of the islet show no [Ca²⁺]_i influx (right). (D) Mean ± SE ChR2-activated [Ca²⁺] resulting from quadrant activation regions, presented in rank order, normalized to islet size. (E) Mean ± SE intraislet variation in ChR2-activated [Ca²⁺] relative to the islet average. The relative area of ChR2-activated [Ca²⁺]_i over each quadrant (as in D) was expressed relative to the mean ChR2-activated [Ca²⁺]_i of each islet, and sorted from least to most. (F) Activation of smaller single-cell regions within islets similar to (C). [Ca²⁺] is elevated within and outside the activation region (right). (G) Distribution of the ChR2-activated area upon single-cell activation regions (n = 96 regions). (H) Mean ± SE ChR2-activated area of [Ca²⁺]_i elevation upon varying sizes of activation region. (I) Repeated activation of one quadrant is shown at 0, 10, 20, and 30 min. (J) Mean ± SE intraislet variation in ChR2-activated [Ca²⁺] as in (E), at time 0 and after 30 min of repeated ChR2 stimulation, sorted from least to most based on time-0 measurements. (K) As in (I), for minimum and maximum quadrants at 0, 10, 20, and 30 min during ChR2 stimulation. (L) Mean ± SE ChR2-activated area upon single-cell activation at time 0 and after 10 min of repeated ChR2 stimulation, averaged over the bottom and top 25% areas, and under a second stimulation sorted based on time-0 measurements. Scale bar in (B), (C), and (F) indicates 2% change in fluorescence. Data in (D) and (E) were averaged over n = 25 islets from six mice; data in (G) was averaged over n = 96 regions, 26 islets from five mice. Data in (I–L) were averaged over n = 10 islets from two mice. ^∗∗∗∗ indicates p < 0.0001, ^∗∗ indicates p < 0.01, ^∗ indicates p < 0.05 comparing experimental groups indicated. In (K), maximal values are compared to the minimal values at each time point. Image scale bars, 100 μm. To see this figure in color, go online.