Membrane structure in ultrarapidly frozen, unpretreated, freeze-fractured myocardium

Abstract

Ultrarapid freezing has been applied to monitor the structure of the freeze-fractured myocardial sarcollema. Our two goals were 1) to demonstrate that large areas of membrane can be preserved free of visible ice crystal damage and, thus, be amenable to quantitative analysis and 2) to compare the structure of directly frozen myocardial membranes with conventionally prepared tissue. The E face was most affected by lack of chemical pretreatment. First, our laboratory reported an increase in E face particle density from 379 ± 30/μm2 in conventional fixed tissue to 489 ± 18/μm2 in unpretreated tissue. Discrete arrays of 12-15 nm particles on the E face were a striking feature of the unfixed sarcolemma. However, P face intramembrane particle (IMP) density remained unchanged from previous estimates in fixed tissue. Specialized regions of the sarcolemma were enhanced in ultrarapidly frozen tissue. Particle domains of the adherens junctions were very prominent in forming a cap alongside the gap junctions. Both the P and E faces of the gap junctions were highly ordered into hexagonal arrays. Caveolae in the membrane were infrequent in both P and E faces.