A Parallel Perifusion Slide From Glass for the Functional and Morphological Analysis of Pancreatic Islets

Abstract

An islet-on-chip system in the form of a completely transparent microscope slide optically accessible from both sides was developed. It is made from laser-structured borosilicate glass and enables the parallel perifusion of five microchannels, each containing one islet precisely immobilized in a pyramidal well. The islets can be in inserted via separate loading windows above each pyramidal well. This design enables a gentle, fast and targeted insertion of the islets and a reliable retention in the well while at the same time permitting a sufficiently fast exchange of the media. In addition to the measurement of the hormone content in the fractionated efflux, parallel live cell imaging of the islet is possible. By programmable movement of the microscopic stage imaging of five wells can be performed. The current chip design ensures sufficient time resolution to characterize typical parameters of stimulus-secretion coupling. This was demonstrated by measuring the reaction of the islets to stimulation by glucose and potassium depolarization. After the perifusion experiment islets can be removed for further analysis. The live-dead assay of the removed islets confirmed that the process of insertion and removal was not detrimental to islet structure and viability. In conclusion, the present islet-on-chip design permits the practical implementation of parallel perifusion experiments on a single and easy to load glass slide. For each immobilized islet the correlation between secretion, signal transduction and morphology is possible. The slide concept allows the scale-up to even higher degrees of parallelization.

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“…Schulze et al developed an islet-on-a-chip device fabricated in glass to simultaneously measure NAD(P)H, FAD and Ca 2+ -influx from 5 individual islets. 85 The device had 5 channels running in parallel with separate inlets and outlets (Fig. 7C, left-top).…”

What Is the Metabolic Amplification of Insulin Secretion and Is It (Still) Relevant?

“…In this context, it is worth mentioning that Ca 2+ influx does not simply constitute the signal for granule fusion but that it may affect granule transport and thus the availability of fusion-ready granules. In fact, the biphasic increase in the cytosolic Ca 2+ concentration, often seen in intact islets [100,101], suggests a role for the Ca 2+ signal beyond triggering.…”

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“…Microfluidic devices offer near infinite diversity in design, limited only by the imagination and fabrication skill of the creator, and can be tailored to fit the application or biological question at hand [11] . Microfluidic devices can be fabricated by a wide variety of methods, most of which require specialized equipment and skills such as photolithography [12] , CNC milling [13] , or laser micromaching [14] . The alternative, commercial solutions, whether it be custom or off-the-shelf can be cost-prohibitive, depending on complexity and specialization.…”