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Blebbistatin, a novel inhibitor of myosin II ATPase activity, increases aqueous humor outflow facility in perfused enucleated porcine eyes.

Publication ,  Journal Article
Zhang, M; Rao, PV
Published in: Invest Ophthalmol Vis Sci
November 2005

PURPOSE: To investigate the specific role of myosin II, a critical biochemical determinant of cellular contraction, in modulation of aqueous humor outflow facility through the trabecular meshwork (TM) pathway. METHODS: Expression of the nonmuscle myosin II heavy chains (IIA, IIB, and IIC) in human TM and ciliary body (CB) cells was determined by RT-PCR analyses. The effects of inhibition of myosin II on cell morphology, actomyosin organization, and cell adhesions were evaluated in porcine TM and CB cells treated with blebbistatin, a cell-permeable, specific inhibitor of myosin II adenosine triphosphatase (ATPase) activity. Changes in aqueous humor outflow facility were determined in enucleated porcine eyes by using a constant-pressure Grant perfusion model system. Ultrastructural integrity of the outflow pathway in drug-perfused eyes was analyzed by transmission electron microscopy. RESULTS: Expression of nonmuscle myosin IIA and IIB was confirmed in both human TM and CB cells. Confluent cultures of primary porcine TM and CB cells treated with blebbistatin in the presence of serum revealed dose (10-200 microM)-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 withdrawal from the cell culture media. Blebbistatin did not affect the status of myosin light chain phosphorylation in TM cells. Perfusion of enucleated porcine eyes for 5 hours with 100 and 200 microM blebbistatin produced a significant increase (P < 0.01, n = 7) in aqueous outflow facility (53% and 64%, respectively) from the baseline facility, compared with a 21% facility increase in sham control specimens. The integrity of the inner wall of aqueous plexi in drug-perfused porcine eyes was found to be intact, and TM cell morphology appeared to be similar to that noted in sham-treated eyes. CONCLUSIONS: These data demonstrate that selective inhibition of myosin II in the aqueous humor outflow pathway leads to increased aqueous outflow facility, suggesting a critical role for myosin II in the regulation of aqueous humor outflow facility. This study also suggests myosin II as a potential therapeutic target for lowering intraocular pressure in patients with glaucoma.

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Published In

Invest Ophthalmol Vis Sci

DOI

ISSN

0146-0404

Publication Date

November 2005

Volume

46

Issue

11

Start / End Page

4130 / 4138

Location

United States

Related Subject Headings

  • Trabecular Meshwork
  • Swine
  • Reverse Transcriptase Polymerase Chain Reaction
  • Phosphorylation
  • Perfusion
  • Ophthalmology & Optometry
  • Nonmuscle Myosin Type IIB
  • Nonmuscle Myosin Type IIA
  • Heterocyclic Compounds, 4 or More Rings
  • Eye Enucleation
 

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Zhang, M., & Rao, P. V. (2005). Blebbistatin, a novel inhibitor of myosin II ATPase activity, increases aqueous humor outflow facility in perfused enucleated porcine eyes. Invest Ophthalmol Vis Sci, 46(11), 4130–4138. https://doi.org/10.1167/iovs.05-0164
Zhang, Min, and P Vasantha Rao. “Blebbistatin, a novel inhibitor of myosin II ATPase activity, increases aqueous humor outflow facility in perfused enucleated porcine eyes.Invest Ophthalmol Vis Sci 46, no. 11 (November 2005): 4130–38. https://doi.org/10.1167/iovs.05-0164.
Zhang, Min, and P. Vasantha Rao. “Blebbistatin, a novel inhibitor of myosin II ATPase activity, increases aqueous humor outflow facility in perfused enucleated porcine eyes.Invest Ophthalmol Vis Sci, vol. 46, no. 11, Nov. 2005, pp. 4130–38. Pubmed, doi:10.1167/iovs.05-0164.

Published In

Invest Ophthalmol Vis Sci

DOI

ISSN

0146-0404

Publication Date

November 2005

Volume

46

Issue

11

Start / End Page

4130 / 4138

Location

United States

Related Subject Headings

  • Trabecular Meshwork
  • Swine
  • Reverse Transcriptase Polymerase Chain Reaction
  • Phosphorylation
  • Perfusion
  • Ophthalmology & Optometry
  • Nonmuscle Myosin Type IIB
  • Nonmuscle Myosin Type IIA
  • Heterocyclic Compounds, 4 or More Rings
  • Eye Enucleation