A new view of the human trabecular meshwork using quick-freeze, deep-etch electron microscopy

Area of Science:

• Ophthalmology
• Cell Biology
• Biophysics

Background:

• Conventional transmission electron microscopy (TEM) of the juxtacanalicular tissue (JCT) shows large flow passages, questioning its role in outflow resistance.
• The extracellular matrix (ECM) preservation in the trabecular meshwork (TM) is crucial for understanding aqueous humor dynamics.

Purpose of the Study:

• To investigate if the extracellular matrix fills apparent open spaces in the JCT using quick-freeze/deep-etch (QF/DE) TEM.
• To compare QF/DE ultrastructural preservation with conventional TEM for the TM and inner wall of Schlemm's canal (SC).

Main Methods:

• Human eyes were fixed under varying pressures (15 or 45 mmHg).
• Trabecular meshwork (TM) and inner wall of Schlemm's canal (SC) were examined using QF/DE and conventional TEM.
• Ultrastructural details of the ECM and basement membrane were analyzed.

Main Results:

• QF/DE provided superior 3D ultrastructural detail of the TM compared to conventional TEM.
• More ECM was observed in the JCT with QF/DE, but micron-sized open spaces remained.
• The basal lamina of the SC inner wall was more complex with QF/DE and showed reduced continuity at higher pressures.

Conclusions:

• Despite revealing more ECM, QF/DE still shows open spaces in the JCT, questioning its contribution to outflow resistance.
• The discontinuous basal lamina of the SC inner wall may be pressure-induced, related to giant vacuole formation, rather than a normal feature.
• QF/DE offers advantages for ultrastructural study of the TM, though further refinement may be needed to fully characterize the JCT ECM.