Micromanipulation of retinal neurons by optical tweezers

Micromanipulation by optical tweezers has been tested in cultures of mature isolated retinal cells to determine its potential for use in creating synaptic circuits in vitro. Rod and cone photoreceptors as well as other retinal nerve cell types could be optically trapped with a 980 nm diode laser mounted on an inverted light microscope using a 40x oil immersion objective numerical aperture of 1.3. Manipulation was done under sterile conditions using transparent culture dishes. To form cell groups, one half of a culture dish was made less adhesive by application of a thin layer of silicone elastomer. Unattached cells were trapped and relocated next to cells lying on an adhesive culture substrate. Optical trapping did not affect the ability of neurons to subsequently attach to the culture substrate. Up to 60% of trapped cells survived for 2 or more days. The pattern and rate of process outgrowth for manipulated cells was comparable to unmanipulated cells and by 2 days, cell-cell contacts were observed. Cultures were fixed at 2 and 5 days for electron microscopy. Organelle, nuclear and cytoplasmic structure of manipulated cells was completely normal and in photoreceptors, synaptic vesicles and ribbons were intact. Optical tweezers, therefore, provide a benign technique with which to micromanipulate whole neurons. The procedures also bestow increased precision to the study of cell-cell interactions by allowing the selection of potentially interacting cell types at a single cell level.