ISSN 1470-3947 (print) | ISSN 1479-6848 (online)

Endocrine Abstracts (2018) 59 OC3.2 | DOI: 10.1530/endoabs.59.OC3.2

Glucose regulates pancreatic [beta] cell Ca2+ dynamics and connectivity in vivo in the anterior chamber of the mouse eye

Victoria Salem1, Kinga Suba1, Aldara Martin-Alonso1, Luis Fernando Delgadillo Silva2, Nadeem Akhtar2, Neda Mousavy1, Eleni Georgiadou1, David Gaboriau1, Stephen Rothery1, Theodorus Stylianides3, Piero Marchett4, Linford Briant5, Nikolay Ninov2, David Hodson6, Walter Distaso1 & Guy Rutter1

1Imperial College London, London, UK; 2Technische Universität Dresden, Dresden, Germany; 3Loughborough University, Loughborough, UK; 4University of Pisa, Pisa, Italy; 5University of Oxford, Oxford, UK; 6University of Birmingham, Birmingham, UK.

Background and Aims: β cell connectivity is a feature of pancreatic islets in vitro but its existence in vivo, when innervated and continuously perfused with blood, has not yet been demonstrated. We imaged islets engrafted in the anterior chamber of the mouse eye (ACE) to explore this question.

Methods: Mouse (C57BL6, Ins1Cre::GCaM6mf/f) or human islets infected with adenovirus to express GCaMP6m, were engrafted and Ca2+ imaging performed under anaesthesia. Glucose or insulin were administered intravenously to achieve low glucose (4–6 mM) or high glucose (25–30 mM) conditions Data were collected on a spinning disc confocal microscope using a 20×, 1.0 NA water immersion objective (3 Hz). Following movement correction, Ca2+ traces were analyzed with Image J. Connectivity analysis was performed with custom-built scripts in Matlab.

Results: Ca2+ waves spreading across the islet in 5/5 animals were observed. Even at low glucose concentrations, β cells form a highly connected syncytium. Increasing glucose concentrations augmented the proportion of connected β cells from 65 to 86% (n=5; P=0.02) and correlation strength (Pearson R with bootstrapping) from 0.34±0.07 to 0.46±0.08 (n=5; P=0.05). Granger causality analysis indicated that cells which responded first during Ca2+ pulses were causally linked to the activity of the largest number of other β cells in the islet. Moreover, the presence of a super-connected β cell subpopulation (8.7±3.6% of cells) was revealed by signal binarisation and Monte Carlo randomization. Pearson connectivity was increased from 58.3% to 63.9% (n=1 animal) in engrafted human islets

Conclusions: We demonstrate intercellular connectivity between β cells within the islet in vivo under conditions of normal islet perfusion and innervation. These findings are consistent with the existence of islet pacemaker cells which coordinate Ca2+ dynamics and possibly pulsatile insulin secretion in the physiological setting.