Decreased amplitude in intracellular Ca²⁺ responses to tolbutamide in <i>Lepr^flox/flox RIP-Cre</i> adult islets.

<p>A: Representative recordings from a control <i>Lepr^flox/flox</i> islet (solid line) and a <i>Lepr^flox/flox RIP-Cre</i> islet (dotted line) in response to tolbutamide. B: Graph plotting ΔFmax-Fmin of each peak in response to tolbutamide in the population of islets that showed two peaks. Data are expressed as mean ± SEM. Statistical analysis was performed using Student t test, *** p<0.0001. Responses are representative of 22 islets from 4 mice per group.</p

Transmission electron microscopy reveals autophagy within <i>Lepr^flox/flox RIP-Cre</i> β-cells.

<p>Pancreas sections from <i>Lepr^flox/flox</i> (A) and <i>Lepr^flox/flox RIP-Cre</i> (B and D) mice were analyzed by transmission electron microscopy (magnification 9300X and 11000X) and quantified (C). Multigranular bodies were numerous in <i>Lepr^flox/flox RIP-Cre</i> β-cells compared to <i>Lepr^flox/flox</i> β-cells (white squares). Events of macroautophagy (<a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0071075#pone-0071075-g004" target="_blank">Figure 4D</a>, upper inset) and microautophagy (<a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0071075#pone-0071075-g004" target="_blank">Figure 4D</a>, bottom inset) were captured in <i>Lepr^flox/flox RIP-Cre</i> β-cells. Scale bar = 2 µm (A and B) and 0.5 µm (D). Micrographs are representative of 3 pancreata analyzed per group. Data are expressed as mean ± SEM. Statistical analysis was performed using Student t test, ** p<0.01.</p

<i>Lepr^+/+ RIP-Cre</i> mice present impaired β-cell Ca²⁺ signaling in response to glucose and tolbutamide.

<p>A: Representative [Ca²⁺]_i recordings of a <i>Lepr^+/+</i> islet (left panel) and <i>Lepr^+/+ RIP-Cre</i> islet (right panel) in response to increasing glucose (G) concentrations and potassium chloride (KCl) from 5–6 week old mice. B: Representative [Ca²⁺]_i recordings of a <i>Lepr^+/+</i> islet (solid line) and <i>Lepr^+/+ RIP-Cre</i> islet (dotted line) in response to tolbutamide. C: Graph plotting the AUC of the Ca²⁺ transients during the stimulation with tolbutamide. Data are expressed as mean ± SEM. Statistical analysis was performed using Student t test. *** p<0.0001. Graphs are representative of 18–20 islets from 3 mice per group (in response to glucose) and 16–27 islets from 2–3 mice per group (in response to tolbutamide).</p

<i>Lepr^flox/flox RIP-Cre</i> pancreatic β-cells display impaired intracellular Ca²⁺ oscillations in response to glucose.

<p>A: [Ca²⁺]_i recordings of a <i>Lepr^flox/flox</i> islet (left panel) and <i>Lepr^flox/flox RIP-Cre</i> islet (right panel) in response to increasing glucose (G) concentrations and potassium chloride (KCl) from adult mice. B and C: Representative [Ca²⁺]_i recordings showing three different regions per islet of a <i>Lepr^flox/flox</i> islet (left panel) and a <i>Lepr^flox/flox RIP-Cre</i> islet (right panel) from adult (B) and neonatal (C) mice. Graphs are representative of 17–20 islets from 3 neonatal mice per group, and 37–38 islets from 3–4 adult mice per group.</p

Similar glucose metabolic rates in islets from <i>Lepr^flox/flox</i> and <i>Lepr^flox/flox RIP-Cre</i> adult mice.

<p>A: Two [NAD(P)H]_i representative recordings of a <i>Lepr^flox/flox</i> islet (solid line) and a <i>Lepr^flox/flox RIP-Cre</i> islet (dotted line) in response to glucose (G) and sodium azide (NaN₃). B: Graph plotting percentage of AUC/min in response to different glucose concentrations and normalized to the maximum reduction level obtained with 3 mM NaN₃. Data are expressed as mean ± SEM. Statistical analysis was performed using Student t test. Graphs are representative of 32–34 islets from 3 mice per group.</p