Dysfunction of Anoctamin 1 (Ano1) in F508del mouse pancreatic b-cell
(2019) European Cystic Fibrosis Society 16th ECFS Basic Science Conference — Location: Dubrovnik, Croatia (27.March.2019)
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- Cystic Fibrosis (CF) is complicated by a form of diabetes (CFRD) mainly characterized by a gradual decline in insulin secretion. Although not apparent at birth, its prevalence increases with age, complicating CF in about 50% of the patients > 35 years, worsening their condition and increasing mortality. Yet little is known on the underlying mechanisms and the functional defects of CFRD within the β-cell. Interestingly, the Anoctamin 1 (Ano1) channel, a calcium-activated chloride channel, has been shown to contribute to insulin secretion and its activity appears to be linked to CFTR function. First Ano1 is expressed by mouse pancreatic b-cells as observed in rat and human b-cells. By immunohistochemistry we localized Ano1 in pancreatic islets and by patch clamp analysis we detected a Ca2+-activated anion channel. Increase in extracellular glucose is a pacemaker inducing cyclic oscillations in b-cell membrane potential. After an initial depolarization, due to closure of the KATP channels, alternating phases of depolarization (active) and repolarization (silent) phases can be observed partially explained by the gating of Ano1. During the active phases, spikes then repeated bursts of action potentials (AP) occur witnessing that the threshold for opening voltage-dependent Ca2+ channels has been reached, allowing Ca2+ entry and triggering insulin release. By patch clamp, we studied the fluctuations of the b-cell membrane potential induced by glucose, 11.1 mM. Their pattern was similar in young F508del mice (< 1 year) vs WT mice, except a hyperpolarized b cell membrane resting potential. On the contrary this pattern was altered in aged F508del mice (>1 year) when compared to matched WT mice. In these F508del mice, the resting membrane potential was hyperpolarized (-81 vs -74 mV in WT; p=0.0002) and the number of AP/30 min was drastically reduced (811 vs 1394 in WT; p=0.01) with the bursting second phase peak amplitude partially repolarized (-29 vs -15 mV in WT; p=0.009). Moreover the duration of the silent phase separating the first spiking active phase and the second bursting continuous phase was increased (715 vs 275 sec in WT; p=0.025). The Ano1 potentiator bromophenyltetrazolbenzamide (Fact) was used to further study Ano1 contribution, first in WT mice. When Fact was added simultaneously with the increased glucose concentration, no silent phase was anymore observed and the number of AP/30 min was increased from 1394 to 3475 (p=0.0002). When Fact was added after the increase in glucose concentration, the AP events/30 min increased from 1728 to 3240 (p=0.01) while the AP peak was further depolarized (-16 vs -21 mV; p=0.007). Thus in F508del as in WT b-cell, Fact increased AP number and at least partially corrected the decreased electrical activity of F508del b-cells. Insulin secretion by isolated islets is clearly increased by Fact in WT mice. Its effect in aged F508del mice was therefore examined here. In conclusion, b-cell from F508del mice exhibit membrane hyperpolarization and as the mice aged, dysfunctional Ano1 gating appears leading to reduced number of AP/30 min. Fact appears to correct Ano1 dysfunction in F508del mice and thus is of potential interest in CFRD.
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Raphaël Crutzen, Delion, M., Mottais, A., Leal, T., Philippe E. Golstein, Alain Boom, & Renaud Beauwens. (2019). Dysfunction of Anoctamin 1 (Ano1) in F508del mouse pancreatic b-cell. European Cystic Fibrosis Society 16th ECFS Basic Science Conference, Dubrovnik, Croatia. https://hdl.handle.net/2078.5/26177