Imaging of outer segment periodicities in unstained cryoultramicrotomy sections of the frog retina.
Frog retinas were examined following cryoultramicrotomy. The extent of glutaraldehyde cross-linking ranged from maximally 2% for 2 h to no cross-linking. When the tissue was cryoprotected, the effects of hypotonic, isotonic and hypertonic media on outer segment banding periodicities were studied. The specimen was cryosectioned at 193 K to 113 K with sections collected on a dry knife (213 K to 123 K). The sections were freeze-dried and only rehydrated with phosphotungstic acid section stain when transmission electron microscopy was used to establish the extent of gross retinal morphology for each fixation protocol employed. As the extreme of no cross-linking and no cryoprotection was approached and reached, conventional sections which displayed extensive retinal morphology were not obtained. Instead, fragments of retinal tissue resulted which showed periodicities of alternating light and dark bands. These periodicities are interpreted as photoreceptor outer segment discs with contrast formation depending upon an interplay between three factors: mass density differences between disc and interdisc space, section cutting angle, and section thickness.
D000818:Animals; D003451:Cryoprotective Agents; D005404:Fixatives; D005629:Frozen Sections; D005976:Glutaral; D006982:Hypertonic Solutions; D007038:Hypotonic Solutions; D008854:Microscopy, Electron; D008867:Microtomy; D010786:Photoreceptor Cells / Q000648:ultrastructure*; D011894:Rana pipiens; D012374:Rod Cell Outer Segment / Q000648:ultrastructure*
D003451:Cryoprotective Agents; D005404:Fixatives; D006982:Hypertonic Solutions; D007038:Hypotonic Solutions; D005976:Glutaral