May 2005
Volume 46, Issue 13
ARVO Annual Meeting Abstract  |   May 2005
Blebbistatin, a Specific Inhibitor of Myosin II, Increases Aqueous Humor Outflow Facility in Perfused Porcine Eyes
Author Affiliations & Notes
  • M. Zhang
    Duke University School of Medicine, Durham, NC
  • P.V. Rao
    Ophthalmology,Pharmacology and Cancer Biology,
    Duke University School of Medicine, Durham, NC
  • Footnotes
    Commercial Relationships  M. Zhang, None; P.V. Rao, None.
  • Footnotes
    Support  NIH Grants EY012201, EY013573 and Research To Prevent Blindness
Investigative Ophthalmology & Visual Science May 2005, Vol.46, 1351. doi:
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      M. Zhang, P.V. Rao; Blebbistatin, a Specific Inhibitor of Myosin II, Increases Aqueous Humor Outflow Facility in Perfused Porcine Eyes . Invest. Ophthalmol. Vis. Sci. 2005;46(13):1351.

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      © ARVO (1962-2015); The Authors (2016-present)

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Abstract: : Purpose: To investigate the specific role of myosin II in the modulation of aqueous humor outflow facility through the trabecular meshwork pathway. Methods: Expression of non–muscle Myosin II heavy chains (IIA, IIB and IIC) in human trabecular meshwork (TM) and ciliary body (CB) cells was determined by RT–PCR analyses. The effects of blebbistatin, a cell–permeable myosin II ATPase–specific inhibitor, on actomyosin organization and cell adhesions (both focal adhesions and adherens junctions) were evaluated in porcine TM and CB cells, using immunofluorescence techniques. Changes in aqueous humor outflow facility were determined using enucleated porcine eyes and a constant–pressure Grant perfusion model system. Ultrastructural integrity of the outflow pathway in drug–perfused eyes was analyzed by transmission electron microscopy. Results: Both human TM and CB cells were confirmed to express non–muscle myosin IIA and IIB. Confluent cultures of primary porcine TM and CB cells treated with blebbistatin in the presence of serum revealed dose (25–200 µM) dependent changes in cell morphology, decreases in actin stress fiber content, and in focal adhesions and adherens junctions. These changes were found to be reversible within 24 hours of drug wash–out from the cell culture media. Blebbistatin did not have any effects on myosin light chain phosphorylation. Perfusion of enucleated porcine eyes with either 100 µM or 200 µM blebbistatin for 5 hours, produced a dose–dependent and significant increase (P<0.01, n=7) in aqueous outflow facility by 53% and 64%, respectively, from the baseline facility, as compared to a 21% facility increase in sham controls. The integrity of the inner wall of aqueous plexi in drug perfused eyes was found to be intact and TM cell morphology appeared to be normal as well. Conclusions: These data demonstrate that selective inhibition of Myosin II leads to an increase in aqueous outflow facility, thereby suggesting a specific role for Myosin II in the regulation of aqueous outflow facility.

Keywords: outflow: trabecular meshwork • signal transduction: pharmacology/physiology • cytoskeleton 

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