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Accessibility of colloidal gold and horseradish peroxidase to cytosolic spaces in Limulus ventral photoreceptors

Published online by Cambridge University Press:  02 June 2009

L. J. Rubin ,
M. Womble ,
J. E. Brown  and
T. E. Finger
L. J. Rubin*
Affiliation:
Department of Ophthalmology, WashingtonUniversity School of Medicine, St. Louis
M. Womble
Affiliation:
Department of Cellular and Structural Biology, University of Colorado School of Medicine, Denver
J. E. Brown
Affiliation:
Department of Ophthalmology, WashingtonUniversity School of Medicine, St. Louis
T. E. Finger
Affiliation:
Department of Cellular and Structural Biology, University of Colorado School of Medicine, Denver
*
Correspondence to: Leona J. Rubin, Department of Pharmacology, 8103, Washington University School of Medicine, 660 S. Euclid Avenue, St. Louis, MO 63110, USA.
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Abstract

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 postulated 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 arbitrary locus of ventral photoreceptor cells have access to all of the presumed sites of the phototransduction cascade.

Keywords

LimulusColloidal goldHRPPhototransductionIntracellular diffusion
Type
Research Articles
Information
Visual Neuroscience , Volume 2 , Issue 2 , February 1989 , pp. 89 - 96
Copyright
Copyright © Cambridge University Press 1989

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