Accessibility of colloidal gold and horseradish peroxidase to cytosolic spaces in Limulus ventral photoreceptors.
Physiological studies of intracellular messengers frequently employ intracellular injections of large molecules that either monitor or modulate the metabolism of the messenger cascade. Injected molecules have unknown mobility in the cytosol and unknown accessibility to various cytosolic compartments, including those postuiated to be traversed by intracellular messenger molecules. In order to determine whether injected molecules have access to the confined spaces through which messenger molecules must diffuse, we injected 5-nm colloidal gold or horseradish peroxidase, or both, into Limulus ventral photoreceptors. Injections were made by applying pressure pulses to the back of an intracellular micropipette that also monitored membrane voltage. The tissue was fixed at varying times after injection and processed for electron microscopy by conventional techniques. Cells fixed 1-3 min after injection contained HRP reaction product only in the cell body. HRP reaction product was found at varying distances down axons in direct relation to the interval between injection and fixation. Colloidal gold particles were found throughout the cell body but not in axons of tissue fixed 1-3 min after injection. Both HRP reaction product and 5-nm colloidal gold particles were observed within the microvillar projections of internal and external rhabdomere, as well as within the extracisternal spaces of endoplasmic reticulum. We conclude that large molecules injected from an intracellular micropipette into an arbitary locus of ventral photoreceptor cells have access to all of the presumed sites of the phototranduction cascade.
Rubin
L J
LJ
Department of Ophthalmology, Washington University School of Medicine, St. Louis, MO 63110.
Womble
M
M
Brown
J E
JE
Finger
T E
TE
D000818:Animals; D003600:Cytosol / Q000378:metabolism*; D006046:Gold / Q000493:pharmacokinetics*; D006735:Horseradish Peroxidase / Q000493:pharmacokinetics*; D006737:Horseshoe Crabs / Q000378:metabolism*; D007267:Injections; D007425:Intracellular Membranes; D010786:Photoreceptor Cells / Q000378:metabolism* / Q000648:ultrastructure
D006046:Gold; D006735:Horseradish Peroxidase