Hostname: page-component-848d4c4894-ndmmz Total loading time: 0 Render date: 2024-04-30T15:48:20.864Z Has data issue: false hasContentIssue false
  • English
  • Français

Photoreceptor cells dissociated from the compound lateral eye of the horseshoe crab, Limulus polyphemus, I: Structure and ultrastructure

Published online by Cambridge University Press:  02 June 2009

Robert N. Jinks ,
W. J. Brad Hanna ,
George H. Renninger  and
Steven C. Chamberlain
Robert N. Jinks
Affiliation:
Institute for Sensory Research, Syracuse University, Syracuse Department of Bioengineering and Neuroscience, Syracuse University, Syracuse
W. J. Brad Hanna
Affiliation:
Biophysics Interdepartmental Group, Department of Physics, University of Guelph, Guelph
George H. Renninger
Affiliation:
Biophysics Interdepartmental Group, Department of Physics, University of Guelph, Guelph
Steven C. Chamberlain
Affiliation:
Institute for Sensory Research, Syracuse University, Syracuse Department of Bioengineering and Neuroscience, Syracuse University, Syracuse
Get access Rights & Permissions [Opens in a new window]

Abstract

Isolated photoreceptors are desirable for whole-cell and patch-clamp studies of functional properties of visual processes that cannot be clearly analyzed when the photoreceptors are coupled. The retina of the compound lateral eye of the horseshoe crab, Limulus polyphemus, was dissociated into individual retinular cells using an enzyme pretreatment consisting of collagenase, papain, and trypsin, and a two-stage mechanical dissociation. These photoreceptors are functionally viable in an organ culture medium for up to 1 week and possess naked arhabdomeral and rhabdomeral segment membranes which are easily accessible for whole-cell recordings. A dissection technique was also developed whereby the retinal epidermis and neural plexus, as well as the second-order eccentric cells, could be separated from the ommatidia of the compound lateral eye in one simple step, providing viable isolated ommatidia attached to the cornea. The enzyme pretreatment used for dissociating the retina was then used to remove the individual ommatidia from the corneal cones.

Hoffman modulation contrast microscopy was used to develop a reliable method for sorting and collecting viable isolated retinular cells for morphological and electrophysiological studies. Morphological analysis using light microscopy and scanning and transmission electron microscopy revealed that isolated retinular cells are morphologically nearly identical to retinular cells in situ. Isolated retinular cells possess a normal rhabdomere with no apparent loss of microvillar membrane as a result of the isolation process. Ommatidia can presently be isolated with up to six retinular cells possessing essentially normal structure and ultrastructure including thick rays of rhabdom. Isolated ommatidia possess naked A-segment membranes which are also well suited for whole-cell recording techniques.

Keywords

DissociationCollagenaseTrypsinPapainIsolated photoreceptors/ommatidiaRhabdomHoffman microscopy
Type
Research Articles
Information
Visual Neuroscience , Volume 10 , Issue 4 , July 1993 , pp. 597 - 607
Copyright
Copyright © Cambridge University Press 1993

Access options

Get access to the full version of this content by using one of the access options below. (Log in options will check for institutional or personal access. Content may require purchase if you do not have access.)

References

Barlow, R.B. Jr, (1983). Crcadian rhythms in the Limulus visual system. Journal of Neuroscience 3, 856–870.CrossRefGoogle Scholar
Barlow, R.B. Jr, & Kaplan, E. (1971). Limulus lateral eye: Properties of receptor units in the unexcised eye. Science 174, 1027–1029.CrossRefGoogle ScholarPubMed
Barlow, R.B. Jr, & Kaplan, E. (1977). Properties of visual cells in the lateral eye of Limulus in situ. Intracellular recordings. Journal of General Physiology 69, 203–220.CrossRefGoogle ScholarPubMed
Barlow, R.B. Jr, Kaplan, E., Renninger, G.H. & Saito, T. (1987). Circadian rhythms in Limulus photoreceptors. I. Intracellular studies. Journal of General Physiology 89, 353–378.CrossRefGoogle ScholarPubMed
Bayer, D.S. & Barlow, R.B. Jr, (1978). Limulus ventral eye: Physiological properties of photoreceptor cells in organ culture medium. Journal of General Physiology 72, 539–564.CrossRefGoogle ScholarPubMed
Calman, B.G. & Chamberlain, S.C. (1982). Distinct lobes of Limulus ventral photoreceptors. II. Structure and ultrastructure. Journal of General Physiology 80, 839–862.CrossRefGoogle ScholarPubMed
Calman, B.G. (1988). Neuroanatomical studies of the visual system of Limulus polyphemus. Syracuse University, Institute for Sensory Research Special Report ISR-S-26.Google Scholar
Chamberlain, S.C. & Barlow, R.B. Jr, (1984). Transient membrane shedding in Limulus photoreceptors: Control mechanisms under natural lighting. Journal of Neuroscience 4, 2792–2810.CrossRefGoogle ScholarPubMed
Chamberlain, S.C. & Barlow, R.B. Jr, (1987). Control of structural rhythms in the lateral eye of Limulus: Interactions of natural lighting and circadian efferent activity. Journal of Neuroscience 7, 2135–2144.CrossRefGoogle ScholarPubMed
Clark, A.W., Millecchia, R. & Mauro, A. (1969). The ventral photoreceptor cell of Limulus. I. The microanatomy. Journal of General Physiology 54, 289–309.CrossRefGoogle ScholarPubMed
Fahrenbach, W.H. (1968). The morphology of the eyes of Limulus. I. Cornea and epidermis of the compound eye. Zeitschrift für Zellforschung und Mikroskopische Anatomie 87, 278–291.CrossRefGoogle ScholarPubMed
Fahrenbach, W.H. (1969). The morphology of the eyes of Limulus. II. Ommatidia of the compound eye. Zeitschrift für Zellforschung und Mikroskopische Anatomie 93, 451–483.CrossRefGoogle ScholarPubMed
Fahrenbach, W.H. (1975). The visual system of the horseshoe crab Limulus polyphemus International Review of Cytology 41, 285–349.CrossRefGoogle ScholarPubMed
Grenacher, H. (1879). Untersuchungen über das Sehorgan der Arthropoden. Göttingen: Vandenhoek und Ruprecht.Google Scholar
Hanna, B., Jinks, R.N., Zhang, H.-J., Kass, L., Renninger, G.H. & Chamberlain, S.C. (1991). Ultrastructure of Limulus photoreceptors in experimental protocols: Physiologists’ favorite recipes. Society for Neuroscience Abstracts 17, 298.Google Scholar
Hanna, W.J.B., Johnson, E.C., Chaves, D. & Renninger, G.H. (1993). Photoreceptor cells dissociated from the compound lateral eye of the horseshoe crab, Limulus polyphemus. II. Function. Visual Neuroscience 10, 609–620.CrossRefGoogle ScholarPubMed
Hanna, W.J.B. (1992). Electrophysiology of isolated Limulus retinular cells: A patch-clamp study. M.S. Dissertation, University of Guelph.Google Scholar
Kaplan, E. & Barlow, R.B. Jr, (1975). Properties of visual cells in the lateral eye of Limulus in situ. Extracellular recordings. Journal of General Physiology 66, 303–326.CrossRefGoogle Scholar
Kass, L., Pelletier, J.L., Renninger, G.H. & Barlow, R.B. Jr, (1988). Efferent neurotransmission of circadian rhythms in Limulus lateral eye. II. Intracellular recordings in vitro. Journal of Comparative Physiology A 164, 95–105.CrossRefGoogle ScholarPubMed
Kier, C.K. & Chamberlain, S.C. (1990). Dual controls for screening pigment movement in photoreceptors of the Limulus lateral eye: Circadian efferent input and light. Visual Neuroscience 4, 237–255.CrossRefGoogle ScholarPubMed
Lasansky, A. (1967). Cell junctions in ommatidia of Limulus. Journal of Cell Biology 33, 365–383.CrossRefGoogle ScholarPubMed
Millecchia, R., Bradbury, J. & Mauro, A. (1966). Simple photoreceptors in Limulus polyphemus. Science 154, 1199–1201.CrossRefGoogle ScholarPubMed
Millecchia, R. & Mauro, A. (1969 a). The ventral photoreceptor cell of Limulus. II. The basic photoresponse. Journal of General Physiology 54, 310–330.CrossRefGoogle ScholarPubMed
Millecchia, R. & Mauro, A. (1969 b). The ventral photoreceptor cell of Limulus. III. A voltage-clamp study. Journal of General Physiology 54, 331–351.CrossRefGoogle Scholar
Nasi, E. (1991). Electrophysiological properties of isolated photoreceptors from the eye of Lima scabra. Journal of General Physiology 97, 17–34.CrossRefGoogle ScholarPubMed
Nasi, E. & Gomez, M. (1992). Electrophysiological recordings in solitary photoreceptors from the retina of the squid, Loligo pealei. Visual Neuroscience 8, 349–358.CrossRefGoogle ScholarPubMed
Renninger, G.H., Kass, L., Pelletier, J.L. & Schimmel, R. (1988). The eccentric cell of the Limulus lateral eye: Encoder of circadian changes in visual responses. Journal of Comparative Physiology A 163, 259–270.CrossRefGoogle Scholar
Renninger, G.H. (1990). Photoreceptor cells dissociated from the Limulus lateral eye Investigative Ophthalmology and Visual Science (Suppl.) 31, 389.Google Scholar
Smith, T.G., Baumann, F. & Fuortes, M.G.F. (1965). Electrical connections between visual cells in the ommatidium of Limulus. Science 147, 1446–1448.CrossRefGoogle ScholarPubMed
Smith, T.G. & Baumann, F. (1969). The functional organization within the ommatidium of the lateral eye of Limulus. In Mechanisms of Synoptic Transmission (Progress of Brain Research, Vol. 31), ed. Akert, K. & Waser, P.G., pp. 313–349. Amsterdam: Elsevier.CrossRefGoogle Scholar
Trump, B.F., Smuckler, E.A. & Benditt, E.P. (1961). A method for staining epoxy sections for light microscopy. Journal of Ultrastructure Research 5, 343–348.CrossRefGoogle ScholarPubMed
Walrond, J.P. & Szuts, E.Z. (1992). Submicrovillar tubules in distal segments of Squid photoreceptors detected by rapid freezing. Journal of Neuroscience 12, 1490–1501.CrossRefGoogle ScholarPubMed
Warren, M.K. & Pierce, S.K. (1982). Two cell volume regulatory systems in the Limulus myocardium: An interaction of ions and quaternary ammonium compounds. Biological Bulletin 163, 504–516.CrossRefGoogle Scholar
Weiner, W.W. & Chamberlain, S.C. (1991). Morphological properties of the Limulus ommatidial array: Dioptrics and photoreceptors Investigative Ophthalmology and Visual Science (Suppl.) 32, 1128.Google Scholar
Zhang, H.-J., Jinks, R.N., Wishart, A.C., Battelle, B.A., ChamberLain, S.C, Fahrenbach, W.H. & Kass, L. (1993). An enzymatically enhanced recording technique for Limulus ventral photoreceptors: Physiology, biochemistry, and morphology. Visual Neuroscience 10, in press.Google Scholar