The development and validation of ultra-thin-film micro-outlet devices for spatially constraining local O₂ perturbations to capillaries

Kiley, Meghan E (2023) The development and validation of ultra-thin-film micro-outlet devices for spatially constraining local O₂ perturbations to capillaries. Masters thesis, Memorial University of Newfoundland.

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Abstract

Several local mechanisms for oxygen concentration [O₂] sensing and blood flow regulation in the microvasculature have been proposed, but existing evidence fails to account for the sensitivity of vascular responses in specific vessels across the physiological range of tissue [O₂]. We hypothesize that oxygen-mediated blood flow regulation is initiated at the capillary level through oxygen saturation-dependent ATP release from erythrocytes. The purpose of this thesis was to develop and validate a thinfilm micro-outlet device that can impose spatially constrained O₂ perturbations at the capillary level to precisely target capillary level regulation. The device was fabricated using soft lithography techniques and high-precision laser cutting. Devices were lasermachined into polyvinylidene chloride film and spun coat with a 100-micrometer thick layer of polydimethylsiloxane. Rats were anesthetized via intraperitoneal injection of sodium pentobarbital; catheters were introduced into the carotid artery for systemic cardiovascular monitoring and jugular vein for supplemental fluids. The extensor digitorum longus (EDL) muscle was blunt dissected, isolated, and reflected over a microfluidic gas exchange chamber (GEC) mounted in the stage of an inverted microscope. The GEC and EDL were coupled with micro-outlet devices of various designs (diameters: 200, 400, 600, 1000 am). [O₂] in the EDL was dynamically manipulated by imposing [O₂] oscillations while recording intravital video. Our novel composite thin-film micro-outlet devices spatially confined oxygen perturbations to capillaries. Our results demonstrate that our devices can profoundly manipulate capillary SO₂ and simultaneously alter the hemodynamics in vessels directly overlying the micro-outlet without affecting capillary SO₂ at distances greater than 100 μm from the edge of the micro-outlets. All animal protocols were approved by Memorial University’s Institutional Animal Care Committee.

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