A chamber to permit invasive manipulation of adherent cells in laminar flow with minimal disturbance of the flow field.
An obstacle to real-time in vitro measurements of endothelial cell responses to hemodynamic forces is the inaccessibility of the cells to instruments of measurement and manipulation. We have designed a parallel plate laminar flow chamber that permits access to adherent cells during exposure to flow. The "minimally invasive flow device" (MIF device) has longitudinal slits (1 mm wide) cut in the top plate of the chamber to allow insertion of a recording, measurement, or stimulating instrument (e.g., micropipette) into the flow field. Surface tension forces at the slit openings are sufficient to counteract the hydrostatic pressure generated in the chamber and thus prevent overflow. The invasive probe is brought near to the cell surface, makes direct contact with the cell membrane, or enters the cell. The slits provide access to a large number (and choice) of cells. The MIF device can maintain physiological levels of shear stress (<1-15 dyn/cm2) without overflow in the absence and presence of fine instruments such as micropipettes used in electrophysiology, membrane aspiration, and microinjection. Microbead trajectory profiles demonstrated negligible deviations in laminar flow near the surface of target cells in the presence of microscale instruments. Patch-clamp electrophysiological recordings of flow-induced changes in membrane potential were demonstrated. The MIF device offers numerous possibilities to investigate real-time endothelial responses to well-defined flow conditions in vitro including electrophysiology, cell surface mechanical probing, local controlled chemical release, biosensing, microinjection, and amperometric techniques.
Levitan
I
I
Institute for Medicine and Engineering, University of Pennsylvania, Philadelphia 19104-6383, USA.
Helmke
B P
BP
Davies
P F
PF
D000818:Animals; D001698:Biomedical Engineering; D002417:Cattle; D002448:Cell Adhesion; D002478:Cells, Cultured; D004594:Electrophysiology; D004730:Endothelium, Vascular / Q000166:cytology* / Q000502:physiology*; D006439:Hemodynamics; D018056:Hemorheology / Q000295:instrumentation*; D008564:Membrane Potentials; D018408:Patch-Clamp Techniques; D015221:Potassium Channels / Q000378:metabolism
D015221:Potassium Channels