# Multi-dimensional Dynamics of Pancreatic Islet Cells Measured by Image Mapping diSPIM

> **NIH NIH R01** · WASHINGTON UNIVERSITY · 2020 · $406,414

## Abstract

ABSTRACT
Our understanding of cellular dynamics has been advanced significantly by live-cell fluorescence microscopy
experiments. These experiments have yielded discoveries in vesicle trafficking and exocytosis on the
functionally important time and length scales, with specific implications for pancreatic islet function. Live-cell
hyperspectral imaging permits simultaneous measurements of multiple dynamic processes with signal-to-noise
ratios equivalent or superior to filter-based approaches. Currently, the most expedient hyperspectral imaging
systems use confocal microscopy, which is limited by photobleaching and slow imaging speeds. We propose
to develop a novel five-dimensional (x,y,z,t,λ) fluorescence imaging system that provides high spatial,
temporal, and spectral resolution with the minimal possible photobleaching. We will optimize its performance
for investigations of long-standing questions about regulation of insulin secretion. This instrument will combine
two technologies: dual-view Selective-Plane Illumination Microscopy (diSPIM) that yields isotropic diffraction-
limited imaging over extended views in three dimensions, and image mapping spectroscopy (IMS) that permits
whole field hyperspectral detection in a single snapshot. We will build, test, and optimize this novel
instrumentation through two specific aims. Specific aim 1 will focus on building and optimizing a new
hyperspectral IMS system for use with diSPIM, and also adapting software modules for five-dimensional data
acquisition and analysis. To substantiate the advantages of the IMS/diSPIM approach, we will acquire images
simultaneously for at least five biosensor colors with high temporal and spatial resolution. To test and guide
the developments in Aim 1, Specific aim 2 will apply this new instrument to issues in β-cell biology that cannot
be addressed with currently available methods, focusing on two questions of insulin vesicle trafficking and
secretion: a) What is the normal life cycle of an insulin vesicle in the β-cell? Since <10% of the insulin vesicles
are secreted, it has been hypothesized that newly formed vesicles are preferentially secreted, and we propose
that longer-lived vesicles act as a signaling platform. We will use the IMS/diSPIM to measure quantitatively up
to 6 fluorescent probes, which will allow us to track every vesicle in a β-cell as it buds from the Golgi, matures,
and is either secreted or moves, putatively irreversibly, into a long-lived pool. b) Do “readily releasable” and
“reserve” vesicle pools lead to the two phases of glucose-stimulated insulin secretion? The concept of two
pools comes from synaptic vesicle studies, which may differ from the crowded environment of the β-cell, where
first phase secretory events appear to come from vesicles newly arriving at the plasma membrane. We
hypothesize that vesicles in β-cells move along microtubules to sites of exocytosis, and these movements are
regulated by intracellular free calcium activity ([Ca2+...

## Key facts

- **NIH application ID:** 9952124
- **Project number:** 5R01DK115972-03
- **Recipient organization:** WASHINGTON UNIVERSITY
- **Principal Investigator:** David W Piston
- **Activity code:** R01 (R01, R21, SBIR, etc.)
- **Funding institute:** NIH
- **Fiscal year:** 2020
- **Award amount:** $406,414
- **Award type:** 5
- **Project period:** 2018-07-01 → 2022-06-30

## Primary source

NIH RePORTER: https://reporter.nih.gov/project-details/9952124

## Citation

> US National Institutes of Health, RePORTER application 9952124, Multi-dimensional Dynamics of Pancreatic Islet Cells Measured by Image Mapping diSPIM (5R01DK115972-03). Retrieved via AI Analytics 2026-05-22 from https://api.ai-analytics.org/grant/nih/9952124. Licensed CC0.

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