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Lack of cytoplasmic laminated bodies in the lateral geniculate nucleus of the rabbit
Brain Research, 168 (1979) 623-627 © Elsevier/North-Holland Biomedial Press
623
Lack of cytoplasmic laminated bodies in the lateral geniculate nucleus of the rabbit
DAVID L. GLANZMAN, JULIE JOHNS, KAO LIANG CHOW* and LAWRENCE H. MATHERS
Department of Psychology, Stanford University, and Departmentsof Neurology and Structural Biology, Stanford University School of Medicine, Stanford, Calif. 94305 (U.S.A.) (Accepted February 8th, 1979)
The description of 3 functionally distinct classes of neurons (the so-called X-, Y-, and W-cells) in the cat's visual system5,6,7,9,12,t3,2°,21,23 has led to a search for their Structural correlates. Based on parallels between anatomical and physiological data, it has been suggested that the fl, a, and y cells, identified by Boycott and W~issle2 on the basis of Golgi material, might be the anatomical substrates of X-, Y- and W-cells in the retina, and that the Golgi types 2, 1, and 4 of Guillery11 might be the anatomical substrates of X-, Y-, and W-cells in the dorsal lateral geniculate nucleus (LGN)I~, ~a. A major difficulty with these schemes is their dependence on morphological and physiological criteria (e.g., cell body size and axonal conduction velocity) which do not classify cells into discrete categories (cf., e.g., Fig. 9, ref. 2, and Fig. 9, ref. 6). Recent data of LeVay and Ferster 1~, however, suggest the presence of a cytoplasmic laminated body (CLB) 18,22 may serve as a unique morphological marker for LGN X-cells in the cat. The rabbit LGN contains neurons with concentric receptive fields which can be classified as X or Y by means of criteria employed in work on the cat's visual systemt°. (W-type cells have not yet been found in the rabbit LGN, although they are present in the rabbit retina3). But a prior anatomical investigation of the normal rabbit LGN 17 did not report any cells with CLBs. Such cells may have been missed, however, particularly during the light microscopic investigation which involved perfusion with a paraformaldehyde-glutaraldehydemixture and subsequent staining with thionin without prior refixation with osmic acid. Such a method will not reveal the presence of CLBs in the cat's brainS, le. We therefore re-examined the morphology of rabbit geniculate neurons with a method similar to that originally used by Morales, Duncan and Rehmet16. As a control, we also examined cat geniculate neurons using the same method. One adult cat and two adult Dutch-belted rabbits were used. Each animal was anesthetized with Nembutal and then perfused through the left ventricle with a 20 % * To whom all correspondence should be sent.
624 formaldehyde mixture which consisted of 700 ml distilled water, 100 ml Ringer's solution, 93 g sucrose, 1 g chloralhydrate, and 200 ml formalin. The p H of the solution was buffered to 7.4. The perfusions were initiated with one half of the perfusate solution at r o o m temperature, and then completed with the remainder of the perfusate, which had been prechilled to 4 °C 1. Immediately u p o n completion of each perfusion, both L G N s of the animal were dissected out of the brain and cut into blocks o f tissue approximately 2-3 m m 3 in size. These blocks were kept in the primary fixative for 30-rain to 1 hour, and then placed in buffered osmic acid with sucrose 4. The tissue was subsequently dehydrated in graded ethanols and embedded in Araldite 502.
Fig. 1. A: neurons containing CLBs in the cat LGN. Arrows indicate the CLBs. Note that the large cell at the right does not contain a laminated body. Light micrograph of a 1/zm thick section stained with methylene blue. Scale: i 0 ibm. B : electron micrograph of a cat LGN neuron containing two CLBs. Scale: 3/*m.
625 For the light microscopic investigation, the tissue blocks were cut into sections 1 #m thick and stained with methylene blue. The sections were oriented coronally and spanned the entire dorsoventral extent of the LGN. Sections were taken from random parts of each nucleus: no attempt was made to systematically sample the anteriorposterior length of the nucleus. For the electron microscopic investigation, thin sections (70-80 nm) were cut from the tissue blocks with a glass knife on a MT2 Ultramicrotome. These were placed on grids and subsequently stained by immersion of the grids for 30 min in 2 ~ aqueous uranyl acetate and then for 10 min in Reynold's lead citrate m. The sections were viewed with a Siemens Elmiskop 1A. Neurons containing CLBs were a conspicuous feature of our cat L G N material. For example, when every neuron was counted in a 1 # m methylene-blue-stained section, 55, or approximately 10 ~ , of the 571 cells in the section were found to contain at least one CLB. Even though we did not attempt a thorough measurement of CLB frequency in the cat LGN, this estimate is close to that made by Doolin et al. 8. In agreement with prior reportsaA ~, CLBs were found predominately in medium-sized neurons, and tended to occur in clusters (Fig. 1A). Also, there was typically just one CLB per neuron, although an occasional neuron contained two (Fig. 1B). No CLBs were found in our rabbit material although several thousand L G N neurons were carefully examined with light and electron microscopes (Fig. 2). It cannot be definitely excluded that with different procedures some CLBs will be found in the rabbit geniculate. It is also possible, but not likely, that the rabbit geniculate has a very small percentage of CLB-containing neurons which were not detected due to our sampling method. Preliminary physiological data from our laboratory suggest, however, that X-cells represent approximately 2 0 ~ of the total population of geniculate neurons in the rabbit. We therefore conclude that CLBs cannot serve as reliable morphological markers for L G N X-cells in the rabbit. Whether CLBs may so serve in the cat is currently a matter of controversy. Recent data of Kalil and Worden 14 argue against the hypothesized 1~ exclusive correlation between X-cells and CLB-containing neurons. Kalil and Worden's estimate of the number of CLB-containing cells in the cat L G N suggests they are too few to represent the entire population of L G N X-cells (cf. also ref. 8). Interestingly, Kalil and Worden reported that dark rearing results in an increase in the number of cat geniculate neurons containing CLBs. In explanation of this phenomenon it was suggested that dark rearing causes an increased number of X-cells to develop in the L G N at the expense of L G N Y-cells; this functional alteration is reflected structurally in the increased number of CLBs. Rabbit X-cells, like their feline counterparts, have concentrically organized receptive fields, generally give sustained responses to standing contrast stimuli with low and intermediate levels of light adaptation, and exhibit linearity of spatial summation over their receptive fields as determined by the null tesO o. Cat and rabbit X-cells would thus appear to possess quite similar, if not identical, functional organizations. Yet the rabbit L G N does not contain CLBs. It would be interesting to determine whether dark rearing produces an apparent increase in rabbit L G N X-cells
626
Fig. 2. A: neurons in the rabbit LGN. No laminated bodies were seen in these neurons. Light micrograph ofa I/~m thick section stained with methylene blue. Scale: 10/~m. B: electron micrograph of a rabbit LGN neuron. Scale: 1 ibm.
as determined electrophysiologically. The absence of such an effect might indicate a correspondent functional dissimilarity between cat and rabbit X-cells. The authors are indebted to Reed D. Pike for photographic assistance and to Cheryl Joo for secretarial assistance. This research was supported by N I H grants NS 18512 and NS 12151 to K.L.C., by EY 00691 to K.L.C., and EY 02589 to L.H.M. from the National Eye Institute and by a predoctoral fellowship, N I H grant GM 7181 to D.L.G.
627 1 Bodian, D. and Taylor, N., Synapse arising at central node of Ranvier, and note on fixation of the central nervous system, Science, 139 (1963) 330-332. 2 Boycott, B. B. and W~issle, H., The morphological types of ganglion cells of the domestic cat's retina, J. PhysioL (Lond.), 240 (1974) 397-419. 3 Caldwell, J. H. and Daw, N. W., New properties of rabbit retinal ganglion cells, J. Physiol. (Lond.), 276 (1978) 257-276. 4 Caulfield, J. B., Effects of varying the vehicle for OsO4 in tissue fixation, J. Biophysics. Biochem. Cytol., 3 (1957) 827-829. 5 Cleland, B. G., Dubin, M. W. and Levick, W. R., Sustained and transient cells in the cat's retina and lateral geniculate nucleus, J. Physiol. (Lond.), 217 (1971) 473-496. 6 Cleland, B. G. and Levick, W. R., Brisk and sluggish concentrically organized ganglion cells in the cat's retina, J. Physiol. (Lond.), 240 (1974) 421-456. 7 Cleland, B. G., Levick, W. R. and Sanderson, K. J., Properties of sustained and transient ganglion cells in the cat retina, J. Physiol. (Lond.), 228 (1973) 649-680. 8 Doolin, P. F., Barron, K. D. and Kwak, S., Ultrastructural and histochemical analysis of cytoplasmic laminar bodies in lateral geniculate neurons of adult cat, Amer. J. Anat., 121 (1967) 601-622. 9 Enroth-Cugell, C. and Robson, J. G., The contrast sensitivity of retinal ganglion cells of the cat, J. Physiol. (Lond.), 187 (1966) 517-552. 10 Glanzman, D. L. and Miller, L. G., Classification of concentric cells in the rabbit LGN as X or Y based upon linearity of spatial summation, Neurosci. Abstr., 4 (1978) 629. 11 Guillery, R. W., A study of Golgi preparations from the dorsal lateral geniculate nucleus of the adult cat, J. comp. Neurol., 128 (1966) 21-50. 12 Hochstein, S. and Shapley, R. M., Quantitative analysis of retinal ganglion cell classifications, J. Physiol. (Lond.), 262 (1976) 237-264. 13 Hoffmann, K.-P., Stone, J. and Sherman, S. M., Relay of receptive-field properties in dorsal lateral geniculate nucleus of the cat, J. Neurophysiol., 35 (1972) 518-531. 14 Kalil, R. and Worden, I., Cytoplasmic laminated bodies in the lateral geniculate nucleus of normal and dark reared cats, J. comp. Neurol., 178 (1978) 469-486. 15 LeVay, S. and Ferster, D., Relay cell classes in the lateral geniculate nucleus of the cat and the effects of visual deprivation, J. comp. Neurol., 172 (1977) 563-584. 16 Morales, R., Duncan, D. and Rehmet, R., A distinctive laminated cytoplasmic body in the lateral geniculate body neurons of the cat, J. Ultrastr. Res., 10 (1964) 116-123. 17 Ralston, H. J., III and Chow, K. L., Synaptic reorganization in the degenerating lateral geniculate nucleus of the rabbit, J. comp. Neurol., 147 (1973) 321-350. 18 Reynolds, E. S., The use of lead citrate at high pH as an electron-opaque stain in electron microscopy, J. Cell BioL, 17 (1963) 208-212. 19 Sherman, S. M., Hoffmann, K.-P. and Stone, J., Loss of a specific cell type from dorsal lateral geniculate nucleus in visually deprived cats, J. Neurophysiol., 35 (1972) 523-541. 20 Stone, J. and Dreher, B., Projection of X- and Y-cells of the cat's lateral geniculate nucleus to areas 17 and 18 of visual cortex, J. NeurophysioL, 36 (1973) 551-567. 21 Stone, J. and Fukuda, Y., Properties of cat retinal ganglion cells: A comparison of W-cells with Xand Y-cells, J. Neurophysiol., 37 (1974) 722-748. 22 Szlachta, H. L. and Habel, R. E., Inclusions resembling Negri bodies in the brains of non-rabid cats, Cornell Vetinarian, 43 (1953) 207-216. 23 Wilson, P. D., Rowe, M. H. and Stone, J., Properties of relay cells in cat's lateral geniculate nucleus: a comparison of W-cells with X- and Y-cells, J. Neurophysiol., 39 (1976) 1193-1209.
623
Lack of cytoplasmic laminated bodies in the lateral geniculate nucleus of the rabbit
DAVID L. GLANZMAN, JULIE JOHNS, KAO LIANG CHOW* and LAWRENCE H. MATHERS
Department of Psychology, Stanford University, and Departmentsof Neurology and Structural Biology, Stanford University School of Medicine, Stanford, Calif. 94305 (U.S.A.) (Accepted February 8th, 1979)
The description of 3 functionally distinct classes of neurons (the so-called X-, Y-, and W-cells) in the cat's visual system5,6,7,9,12,t3,2°,21,23 has led to a search for their Structural correlates. Based on parallels between anatomical and physiological data, it has been suggested that the fl, a, and y cells, identified by Boycott and W~issle2 on the basis of Golgi material, might be the anatomical substrates of X-, Y- and W-cells in the retina, and that the Golgi types 2, 1, and 4 of Guillery11 might be the anatomical substrates of X-, Y-, and W-cells in the dorsal lateral geniculate nucleus (LGN)I~, ~a. A major difficulty with these schemes is their dependence on morphological and physiological criteria (e.g., cell body size and axonal conduction velocity) which do not classify cells into discrete categories (cf., e.g., Fig. 9, ref. 2, and Fig. 9, ref. 6). Recent data of LeVay and Ferster 1~, however, suggest the presence of a cytoplasmic laminated body (CLB) 18,22 may serve as a unique morphological marker for LGN X-cells in the cat. The rabbit LGN contains neurons with concentric receptive fields which can be classified as X or Y by means of criteria employed in work on the cat's visual systemt°. (W-type cells have not yet been found in the rabbit LGN, although they are present in the rabbit retina3). But a prior anatomical investigation of the normal rabbit LGN 17 did not report any cells with CLBs. Such cells may have been missed, however, particularly during the light microscopic investigation which involved perfusion with a paraformaldehyde-glutaraldehydemixture and subsequent staining with thionin without prior refixation with osmic acid. Such a method will not reveal the presence of CLBs in the cat's brainS, le. We therefore re-examined the morphology of rabbit geniculate neurons with a method similar to that originally used by Morales, Duncan and Rehmet16. As a control, we also examined cat geniculate neurons using the same method. One adult cat and two adult Dutch-belted rabbits were used. Each animal was anesthetized with Nembutal and then perfused through the left ventricle with a 20 % * To whom all correspondence should be sent.
624 formaldehyde mixture which consisted of 700 ml distilled water, 100 ml Ringer's solution, 93 g sucrose, 1 g chloralhydrate, and 200 ml formalin. The p H of the solution was buffered to 7.4. The perfusions were initiated with one half of the perfusate solution at r o o m temperature, and then completed with the remainder of the perfusate, which had been prechilled to 4 °C 1. Immediately u p o n completion of each perfusion, both L G N s of the animal were dissected out of the brain and cut into blocks o f tissue approximately 2-3 m m 3 in size. These blocks were kept in the primary fixative for 30-rain to 1 hour, and then placed in buffered osmic acid with sucrose 4. The tissue was subsequently dehydrated in graded ethanols and embedded in Araldite 502.
Fig. 1. A: neurons containing CLBs in the cat LGN. Arrows indicate the CLBs. Note that the large cell at the right does not contain a laminated body. Light micrograph of a 1/zm thick section stained with methylene blue. Scale: i 0 ibm. B : electron micrograph of a cat LGN neuron containing two CLBs. Scale: 3/*m.
625 For the light microscopic investigation, the tissue blocks were cut into sections 1 #m thick and stained with methylene blue. The sections were oriented coronally and spanned the entire dorsoventral extent of the LGN. Sections were taken from random parts of each nucleus: no attempt was made to systematically sample the anteriorposterior length of the nucleus. For the electron microscopic investigation, thin sections (70-80 nm) were cut from the tissue blocks with a glass knife on a MT2 Ultramicrotome. These were placed on grids and subsequently stained by immersion of the grids for 30 min in 2 ~ aqueous uranyl acetate and then for 10 min in Reynold's lead citrate m. The sections were viewed with a Siemens Elmiskop 1A. Neurons containing CLBs were a conspicuous feature of our cat L G N material. For example, when every neuron was counted in a 1 # m methylene-blue-stained section, 55, or approximately 10 ~ , of the 571 cells in the section were found to contain at least one CLB. Even though we did not attempt a thorough measurement of CLB frequency in the cat LGN, this estimate is close to that made by Doolin et al. 8. In agreement with prior reportsaA ~, CLBs were found predominately in medium-sized neurons, and tended to occur in clusters (Fig. 1A). Also, there was typically just one CLB per neuron, although an occasional neuron contained two (Fig. 1B). No CLBs were found in our rabbit material although several thousand L G N neurons were carefully examined with light and electron microscopes (Fig. 2). It cannot be definitely excluded that with different procedures some CLBs will be found in the rabbit geniculate. It is also possible, but not likely, that the rabbit geniculate has a very small percentage of CLB-containing neurons which were not detected due to our sampling method. Preliminary physiological data from our laboratory suggest, however, that X-cells represent approximately 2 0 ~ of the total population of geniculate neurons in the rabbit. We therefore conclude that CLBs cannot serve as reliable morphological markers for L G N X-cells in the rabbit. Whether CLBs may so serve in the cat is currently a matter of controversy. Recent data of Kalil and Worden 14 argue against the hypothesized 1~ exclusive correlation between X-cells and CLB-containing neurons. Kalil and Worden's estimate of the number of CLB-containing cells in the cat L G N suggests they are too few to represent the entire population of L G N X-cells (cf. also ref. 8). Interestingly, Kalil and Worden reported that dark rearing results in an increase in the number of cat geniculate neurons containing CLBs. In explanation of this phenomenon it was suggested that dark rearing causes an increased number of X-cells to develop in the L G N at the expense of L G N Y-cells; this functional alteration is reflected structurally in the increased number of CLBs. Rabbit X-cells, like their feline counterparts, have concentrically organized receptive fields, generally give sustained responses to standing contrast stimuli with low and intermediate levels of light adaptation, and exhibit linearity of spatial summation over their receptive fields as determined by the null tesO o. Cat and rabbit X-cells would thus appear to possess quite similar, if not identical, functional organizations. Yet the rabbit L G N does not contain CLBs. It would be interesting to determine whether dark rearing produces an apparent increase in rabbit L G N X-cells
626
Fig. 2. A: neurons in the rabbit LGN. No laminated bodies were seen in these neurons. Light micrograph ofa I/~m thick section stained with methylene blue. Scale: 10/~m. B: electron micrograph of a rabbit LGN neuron. Scale: 1 ibm.
as determined electrophysiologically. The absence of such an effect might indicate a correspondent functional dissimilarity between cat and rabbit X-cells. The authors are indebted to Reed D. Pike for photographic assistance and to Cheryl Joo for secretarial assistance. This research was supported by N I H grants NS 18512 and NS 12151 to K.L.C., by EY 00691 to K.L.C., and EY 02589 to L.H.M. from the National Eye Institute and by a predoctoral fellowship, N I H grant GM 7181 to D.L.G.
627 1 Bodian, D. and Taylor, N., Synapse arising at central node of Ranvier, and note on fixation of the central nervous system, Science, 139 (1963) 330-332. 2 Boycott, B. B. and W~issle, H., The morphological types of ganglion cells of the domestic cat's retina, J. PhysioL (Lond.), 240 (1974) 397-419. 3 Caldwell, J. H. and Daw, N. W., New properties of rabbit retinal ganglion cells, J. Physiol. (Lond.), 276 (1978) 257-276. 4 Caulfield, J. B., Effects of varying the vehicle for OsO4 in tissue fixation, J. Biophysics. Biochem. Cytol., 3 (1957) 827-829. 5 Cleland, B. G., Dubin, M. W. and Levick, W. R., Sustained and transient cells in the cat's retina and lateral geniculate nucleus, J. Physiol. (Lond.), 217 (1971) 473-496. 6 Cleland, B. G. and Levick, W. R., Brisk and sluggish concentrically organized ganglion cells in the cat's retina, J. Physiol. (Lond.), 240 (1974) 421-456. 7 Cleland, B. G., Levick, W. R. and Sanderson, K. J., Properties of sustained and transient ganglion cells in the cat retina, J. Physiol. (Lond.), 228 (1973) 649-680. 8 Doolin, P. F., Barron, K. D. and Kwak, S., Ultrastructural and histochemical analysis of cytoplasmic laminar bodies in lateral geniculate neurons of adult cat, Amer. J. Anat., 121 (1967) 601-622. 9 Enroth-Cugell, C. and Robson, J. G., The contrast sensitivity of retinal ganglion cells of the cat, J. Physiol. (Lond.), 187 (1966) 517-552. 10 Glanzman, D. L. and Miller, L. G., Classification of concentric cells in the rabbit LGN as X or Y based upon linearity of spatial summation, Neurosci. Abstr., 4 (1978) 629. 11 Guillery, R. W., A study of Golgi preparations from the dorsal lateral geniculate nucleus of the adult cat, J. comp. Neurol., 128 (1966) 21-50. 12 Hochstein, S. and Shapley, R. M., Quantitative analysis of retinal ganglion cell classifications, J. Physiol. (Lond.), 262 (1976) 237-264. 13 Hoffmann, K.-P., Stone, J. and Sherman, S. M., Relay of receptive-field properties in dorsal lateral geniculate nucleus of the cat, J. Neurophysiol., 35 (1972) 518-531. 14 Kalil, R. and Worden, I., Cytoplasmic laminated bodies in the lateral geniculate nucleus of normal and dark reared cats, J. comp. Neurol., 178 (1978) 469-486. 15 LeVay, S. and Ferster, D., Relay cell classes in the lateral geniculate nucleus of the cat and the effects of visual deprivation, J. comp. Neurol., 172 (1977) 563-584. 16 Morales, R., Duncan, D. and Rehmet, R., A distinctive laminated cytoplasmic body in the lateral geniculate body neurons of the cat, J. Ultrastr. Res., 10 (1964) 116-123. 17 Ralston, H. J., III and Chow, K. L., Synaptic reorganization in the degenerating lateral geniculate nucleus of the rabbit, J. comp. Neurol., 147 (1973) 321-350. 18 Reynolds, E. S., The use of lead citrate at high pH as an electron-opaque stain in electron microscopy, J. Cell BioL, 17 (1963) 208-212. 19 Sherman, S. M., Hoffmann, K.-P. and Stone, J., Loss of a specific cell type from dorsal lateral geniculate nucleus in visually deprived cats, J. Neurophysiol., 35 (1972) 523-541. 20 Stone, J. and Dreher, B., Projection of X- and Y-cells of the cat's lateral geniculate nucleus to areas 17 and 18 of visual cortex, J. NeurophysioL, 36 (1973) 551-567. 21 Stone, J. and Fukuda, Y., Properties of cat retinal ganglion cells: A comparison of W-cells with Xand Y-cells, J. Neurophysiol., 37 (1974) 722-748. 22 Szlachta, H. L. and Habel, R. E., Inclusions resembling Negri bodies in the brains of non-rabid cats, Cornell Vetinarian, 43 (1953) 207-216. 23 Wilson, P. D., Rowe, M. H. and Stone, J., Properties of relay cells in cat's lateral geniculate nucleus: a comparison of W-cells with X- and Y-cells, J. Neurophysiol., 39 (1976) 1193-1209.