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Microscopy & Imaging JoVE (Open Access) Citable · DOI

Microiontophoresis and Micromanipulation for Intravital Fluorescence Imaging of the Microcirculation

DOI: 10.3791/2900-v
What you'll learn
  • ✓ Prepare and calibrate micropipettes for microiontophoresis delivery
  • ✓ Position micropipettes and deliver acetylcholine via electrical current
  • ✓ Measure endothelium-dependent vasodilation using intravital fluorescence imaging
Protocol

Microiontophoresis entails movement of ions from a micropipette in response to a difference in electrical potential between the inside and outside of the micropipette. Biologically active molecules are thereby delivered in proportion to electrical current. We illustrate acetylcholine microiontophoresis in conjunction with micromanipulation to study endothelium-dependent vasodilation in the microcirculation.

Difficulty
advanced
Total time
~2–3 hours per animal preparation and imaging session
Model organism
Not specified (rodent microcirculation assumed)
Biosafety
BSL-1

Steps

1
Prepare and fill micropipettes for microiontophoresis

Fabricate micropipettes and load them with acetylcholine or other biologically active molecules. Calibrate the micropipette resistance and electrical properties before use.

▶ 01:33
2
Set up sample and apply microiontophoresis current

Position the micropipette onto the target arteriole using micromanipulation. Apply electrical current to deliver acetylcholine in proportion to the applied potential difference.

▶ 03:13
3
Image and record fluorescence response in arteriole

Acquire intravital fluorescence images of the microcirculation to measure changes in vascular diameter and endothelial function in response to acetylcholine delivery.

▶ 04:52
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