Development of thin-film micro-outlets for spatially constraining local PO2 perturbations to capillaries in vivo

TL;DR

Researchers developed thin-film micro-outlet devices to precisely control oxygen levels in small muscle regions and observed effects on capillary blood flow.

Contribution

The novel composite thin-film micro-outlet devices enable localized manipulation of oxygen concentration to study microvascular responses in vivo.

Findings

Oxygen oscillations via 400 μm micro-outlets significantly altered capillary RBC oxygen saturation and supply rate.

200 μm micro-outlets also caused significant changes in RBC oxygen saturation and supply rate at low oxygen levels.

The devices spatially confined oxygen perturbations to affect only capillaries directly overlying the outlets.

Abstract

To develop and validate thin-film micro-outlet devices to study microvascular blood flow responses to localized changes in skeletal muscle oxygen concentration ([O2]).

30 male Sprague-Dawley rats (159–194 g) were anesthetized and instrumented to maintain cardiovascular state. The extensor digitorum longus (EDL) muscle was dissected, isolated, and reflected over a gas exchange chamber (GEC) mounted in the stage of an inverted microscope. The GEC and EDL were coupled via a composite, gas permeable membrane, and a gas impermeable film fabricated with laser machined micro-outlets of specific diameters (200, 400, 600, and 1,000 μm). [O2] in the EDL was dynamically manipulated with step-wise oscillations between 7% (1 min) → 12% (1 min) → 2% (1 min) → 7% (1 min), and step challenges from 7% (1 min) → 2% or 12% (2 min), while recording intravital video for capillary RBC oxygen saturation (SO2) and hemodynamic measurements. Oxygen diffusion between tissue and micro-outlet devices was modelled using a finite element mass transport model to further validate experimental results.

[O2] oscillations imposed on capillaries directly overlying 400 μm micro-outlets caused significant changes in RBC SO2 at 12% and 2% [O2], compared to 7% [O2] (p < 0.0001). [O2] oscillations caused significant changes in capillary RBC supply rate (SR) at 2% [O2] versus 7%, and were significantly different at 2% compared to 12% [O2] (p < 0.0014). Similarly, [O2] challenges imposed on capillaries overlying 200 μm micro-outlets also caused significant changes in RBC SO2 at 2% [O2], compared to 7% [O2] (p < 0.0001), and caused significant changes in SR at 2% [O2] compared to 7% (p < 0.0001).

Our composite thin-film devices were fabricated and validated to spatially confine oxygen perturbations to capillaries using micro-outlets of varying diameters. These results demonstrate that our devices can manipulate capillary SO2 and alter capillary RBC SR in vessels directly overlying the micro-outlet without affecting capillary SO2 at a distance from the outlets. Our novel composite thin-film micro-outlet devices demonstrate that capillary blood flow responses can be provoked by manipulating [O2] in tissue regions as small as ∼200 μm in diameter.

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