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Effects on the growth of damaged ganglion cell axons after peripheral nerve transplantation in adult hamsters
168
Brain Research, 377 (1986) 168-172 Elsevier
BRE 21626
Effects on the growth of damaged ganglion cell axons after peripheral nerve transplantation in adult hamsters KWOK-FAI SO 1, YUE-MEI XIAO 2 and YUE-CHENG DIAO 2
1Department of Anatomy, Faculty of Medicine, Universityof Hong Kong (Hong Kong) and 2Institute of Biophysics, Academia Sinica, Beifing (People's Republic of China) (Accepted March 4th, 1986)
Key words: retina - - peripheral nervous system (PNS) nerve transplant - - central nervous system (CNS) regeneration - - hamster
After transplantation of autologous sciatic nerve segments into the retina of adult hamsters for 1-2 months, retrograde labelling with horseradish peroxidase demonstrated a population of ganglion cells situated peripheral to the graft. If an additional lesion was placed between the insertion of the graft and the optic disc at the same time as transplantation, in addition to labelled cells situated peripheral to the graft, retrograde labelling with horseradish peroxidase demonstrated a population of labelled neurons located between the graft and the optic disc which was not observed in animals without the additional lesion. Since the axons of this population of cells would have to turn around away from their normal course towards the optic disc and travel for about 1.5 mm in order to grow into the graft, it suggests that the peripheral nerve graft might play an active role in attracting and/or guiding damaged ganglion cell axons to grow into it.
Recent experiments using adult rodents have demonstrated that d a m a g e d retinal ganglion cell axons can regenerate for a long distance into a segment of autologous peripheral nerve graft inserted into the eye of rats 9 or hamsters t~. Limited growth of such axons has also been observed when the graft was placed into the optic tract of hamsters 1°. All these experiments agree with the suggestion that the peripheral nervous system (PNS) environment is favourable for supporting extensive regrowth of axons in the adult mammalian CNS 1. H o w e v e r , it is not clear how the d a m a g e d ganglion cell axons grew into the graft in the first place. It is possible to suggest that the graft might play an active role in attracting and/or guiding the regenerating axons to grow into it. Or the graft might play a passive role in the sense that it intercepts cut ganglion cell axons located peripheral to the graft which are trying to grow towards the direction of the optic disc. It is, however, difficult to differentiate between these two possibilities in the previous experiments 9'n. In o r d e r to investigate this p r o b l e m further, we have studied the response of ganglion cells whose axons would not normally be intercepted by
the graft. The results suggest that the graft might play an active role in attracting and/or guiding d a m a g e d ganglion cell axons to grow into it. Thirteen young adult golden hamsters (Mesocricetus auratus) were used for the experiments of transplantation of a segment of autologous sciatic nerve into the retina. Animals with PNS graft alone. In 3 hamsters, one end of the PNS graft was inserted into the superior temporal q u a d r a n t of the left eye about 2 mm from the limbus (Figs. 1 and 2A). O n e - t w o months later, horseradish peroxidase ( H R P ) was applied to the other end of the graft which was laid over the skull and the retinae were processed for H R P histochemistry 4. The details of these procedures have been described previously 9.
Animals with PNS graft plus an additional lesion in the retina. In 10 hamsters, a segment of sciatic nerve was inserted into the superior t e m p o r a l retina about 2 m m from the limbus as described before 9. In 7 of them an additional lesion was placed, at the same time as the transplantation of the graft, in the retina between the location of the insertion of the graft and
Correspondence: K.-F. So, Department of Anatomy, Faculty of Medicine, University of Hong Kong, Hong Kong. 0006-8993/86/$03.50 O 1986 Elsevier Science Publishers B.V. (Biomedical Division)
169
Fig. 1. Photograph of an eye from a hamster showing the optic nerve (ON) and the graft. The star (*) denotes the location where the graft is inserted into the retina.
the optic disc with the tip of a 22-gauge needle (Fig. 2C). One to two months later, H R P retrograde labelling experiments were conducted in 5 of these animals and the retinae were processed for H R P histochemical reaction as described above. In two animals, double fluorescent dyes labelling experiments were performed. The cytoplasmic label True Blue (2%) was applied to the cut end of the graft and 6 days later the cell nucleus label Nuclear Yellow (2%) was applied to the surgically exposed and transected contralateral (n = 1) or ipsilateral (n = 1) optic tract s (Fig. 3A). Two days later, the animals were perfused with cold normal saline. Both retinae were removed, fixed in 10% formol-saline solution for one hour, fiatmounted, dried, coated with DPX, coverslipped and examined with a Zeiss fluorescent microscope. In the remaining 3 animals with a graft in the retina, an additional lesion was placed in the ventral retina ca. 3 mm away from the graft insertion (Fig. 4). One to two months later, H R P was applied to the graft and the retinae were processed for H R P reaction. The transplanted PNS nerve was found to be well attached to the eye (Fig. 1) and subcutaneous tissues overlying the skull after a postgrafting period of 1-2 months. Animals with peripheral nerve transplantation alone. Retrograde H R P labelling experiments were carried out in animals with a segment of PNS nerve inserted into the left retina. H R P applied to the graft
(Fig. 2A) demonstrated, similar to the results obtained from the rat 9, a population of labelled ganglion cells of different sizes situated peripheral to the insertion of the graft in the fiat-mounted retina (Fig. 2B). No cell was observed in the area between the graft and the optic disc and only a few cells were found on the temporal side of the graft (Figs. 2B and 4). Since it is known that the axons would have to be damaged before they would grow into the graft 9'11, it seems to suggest that, similar to the results observed in the cat retina 5, very few ganglion cell axons follow a curved course to reach the optic disc. No labelled neuron was detected in the control, right, retinae. Animals with peripheral nerve transplantation plus an additional lesion. In 7 animals, an additional damage was placed between the insertion of the graft and the optic disc at the same time of transplantation. When H R P was applied to the graft of 5 of these animals (Fig. 2C), in addition to labelled cells situated peripheral to the graft (between 11 and 276 cells, mean + S.D. = 100 + 103), a population of labelled neurons was observed (between 3 and 140 cells, mean + S.D. = 39 + 58) between the graft and the optic disc (Fig. 2D) which was not observed in animals without the additional lesion. A double fluorescent dyes experiment was conducted in the remaining two animals to investigate the origins of the fibers in the graft from the population of cells located between the graft and site of the additional lesion. The experiment was designed to find out whether they were regenerative outgrowth from axotomized neurons or collateral sprouts from uninjured neurons which still retain their axons in the optic tract. If the second possibility is valid, double labelled neurons should be observed after application of True Blue to the graft and Nuclear Yellow to the contralateral (cell B in Fig. 3A) or ipsilateral optic tract. No such cell was observed in the two animals studied suggesting that the axons in the graft originated from neurons whose axons have been damaged. This conclusion is similar to that obtained for the population of regenerating ganglion cells situated peripheral to the graft 9A1. In 3 additional animals, an additional lesion was placed in the ventral retina, ca. 3 mm from the insertion of the graft. After a postgrafting period of 1-2 months, H R P was applied to the graft and labelled neurons were observed peripheral to the graft but no-
170
B
A graft
HRP
T
eye
OC
contra OT
g
I1 HRP
Fig. 2. A: schematic diagram illustrating retrograde HRP labelling experiment in animals with a segment of sciatic nerve transplanted to the left eye. ON, optic nerve; OC, optic chiasm; contra OT, contralateral optic tract. B: diagram of retinal whole mount of a hamster from an experiment (HRR-4, 44 days postgrafting survival time) illustrated in A. Black dots are HRP labelled neurons; g, graft; U, upper retina; T, temporal retinal; x, optic disc. Scale bar, 1 mm. C: schematic diagram illustrating retrograde HRP labelling experiment in animal with PNS graft plus an additional damage (black bar) in the retina. D: diagram of a retinal whole mount of a hamster from an experiment (DOHRR-5, 30 days post grafting survival time) illustrated in C. d, additional damage in the retina. Other abbreviations, same as in B. Note that HRP labelled neurons (black dots) are also found in areas of the retina between the graft and optic disc. Scale bar, 1 mm.
where else (Fig. 4). In all of the above experiments, no labelled n e u r o n was observed in the control, right, retinae. The results from the present experiments in adult hamsters with a graft and an additional lesion in the retina demonstrated that some cut axons which are not in direct apposition with the graft can also regrow into the graft. This was shown by the labelling of a population of n e u r o n s located between the insertion
of the graft and the site of the additional lesion in the retina. Since the axons of this population of cells would have to turn around away from its normal course towards the optic disc, and travel for ca. 1.5 mm from the site of the additional lesion in order to grow into the graft, it suggests that the grafted peripheral nerve might play an active role in attracting and/or guiding damaged ganglion cell axons to grow into it.
171
B
u
A TB
T
Fig. 3. A: schematic diagram illustrating the double fluorescent dyes experiment in animals with PNS graft plus an additional damage (black bar) in the retina. True Blue (TB) was applied to the graft and Nuclear Yellow (NY) to the optic tract in order to trace, in areas between the graft and optic disc, any neuron which has projections in the graft and optic tract (cell B). Cell A is a cell which sends an axon into the graft alone. Cell C is an uninjured neuron which retains its axon in the optic tract. B: diagram of a retinal whole mount of a hamster from an experiment (DOHRR-9, 55 days post grafting survival time) illustrated in A. True Blue-labelled cells (black dots) were located in a fan shaped area peripheral and central to the graft. Nuclear Yellow labelled cells (not illustrated) were located in the remaining part of the retina. Abbreviations as in Fig. 2. Scale bar, 1 mm.
After discrete damage in the retina of adult mouse with no grafting of peripheral nerve, the damaged ax-
ons have b e e n shown to regrow within the retina for a distance of only up to 500 p m 2 or ca. 450 p m 3. But with the addition of the peripheral nerve in the hamster, the damaged ganglion cell axons seem to be able to regrow for a much longer distance within the retina in order to grow into the graft. The mechanisms underlying the attracting and/or
T
Fig. 4. Diagram of a retinal whole mount from an animal (FDHRR-2, 38 days post grafting survival time) with a PNS graft and an additional damage in the ventral retina. Abbreviations as in Fig. 2. Scale bar, 1 mm.
guiding role of the grafted peripheral nerve are not clear at the m o m e n t . The PNS nerve could secrete a diffusable chemical into the e n v i r o n m e n t to attract the damaged axons. For example, denervated peripheral nerves have been shown to secrete nerve growth factor-like substance 6 and nerve growth associated protein 7. O n the other hand, it is also possible that Schwann cells and/or other types of cells might migrate from the graft into the retina thus providing a substrate to guide the axons to grow into the graft. For example, the migration of Schwann cells, fibroblasts and endothelial cells from cut ends of peripheral nerve into a silicone chamber have b e e n observed to precede that of regenerating axons 12. Both of these mechanisms are, however, not mutually exclusive of each other. But if they operate in the hamster, they seem to be effective in attracting and/or guiding
172 d a m a g e d axons only within a certain r a n g e of dis-
This investigation was s u p p o r t e d by the C r o u c h e r
tance, b e c a u s e axons which h a v e b e e n d a m a g e d ca. 3
F o u n d a t i o n of H o n g K o n g . W e t h a n k Mr. K . C . L a u
m m away f r o m the graft could n o t be s h o w n to g r o w
for technical assistance and Miss S. C h u for typing
into the graft.
the manuscript.
1 Aguayo, A.J., Axonal regeneration from injured neurons in the adult mammalian central nervous system. In C.W. Cotman (Ed.), Synaptic Plasticity, The Guilford Press, New York, 1985, pp. 457-484. 2 Goldberg, S. and Frank, B., Will central nervous systems in the adult mammal regenerate after bypassing a lesion? A study in the mouse and chick visual systems, Exp. Neurol., 70 (1980) 675-689. 3 McConnell, P. and Berry, M., Regeneration of ganglion cell axons in the adult mouse retina, Brain Research, 241 (1982) 362-365. 4 Mesulam, M.-M., Tetramethyl benzidine for horseradish peroxidase neurohistochemistry: a non-carcinogenic blue reaction-product with superior sensitivity for visualizing neural afferents and efferents, J. Histochem. Cytochem., 26 (1978) 106-117. 5 Murakami, D., Sesma, M.A. and Rowe, M.H., Characteristics of nasal and temporal retina in Siamese and normally pigmented cats: ganglion cell composition, axon trajectory and laterality of projection, Brain Behav. Evol., 21 (1982) 67-113. 6 Richardson, P.M. and Ebendal, T., Nerve growth activities
in rat peripheral nerve, Brain Research, 246 (1982) 57-64. 7 Skene, P.J.H. and Shooter, E.M., Denervated sheath cells secrete a new protein after nerve injury, Proc. Natl. Acad. Sci. U.S.A., 80 (1983) 4169-4173. 8 So, K.-F. and Schneider, G.E., Abnormal recrossing retinotectal projections after early lesions in Syrian hamster: age-related effects, Brain Research, 147 (1978) 277-295. 9 So, K.-F. and Aguayo, A.J. Lengthy regrowth of cut axons from ganglion cells after peripheral nerve transplantation into the retina of adult rat, Brain Research, 328 (1985) 349-354. 10 Stevenson, J.A., Growth of optic tract axons in nerve grafts in hamsters, Exp. Neurol., 87 (1985) 446-457. 11 Xiao, Y.-M. and So, K.-F., Ganglion cells regenerating axons into peripheral nerve graft transplanted into the retina of adult hamsters, Hong Kong Soc. Neurosci. Abstr., 7 (1985) 13. 12 Williams, L.R., Longo, F.M., Powell, H.C., Lundborg, G. and Varon, S., Spatial-temporal progress of peripheral nerve regeneration within a silicone chamber: parameters for a bioassay, J. Comp. Neurol., 218 (1983) 460-470.
Brain Research, 377 (1986) 168-172 Elsevier
BRE 21626
Effects on the growth of damaged ganglion cell axons after peripheral nerve transplantation in adult hamsters KWOK-FAI SO 1, YUE-MEI XIAO 2 and YUE-CHENG DIAO 2
1Department of Anatomy, Faculty of Medicine, Universityof Hong Kong (Hong Kong) and 2Institute of Biophysics, Academia Sinica, Beifing (People's Republic of China) (Accepted March 4th, 1986)
Key words: retina - - peripheral nervous system (PNS) nerve transplant - - central nervous system (CNS) regeneration - - hamster
After transplantation of autologous sciatic nerve segments into the retina of adult hamsters for 1-2 months, retrograde labelling with horseradish peroxidase demonstrated a population of ganglion cells situated peripheral to the graft. If an additional lesion was placed between the insertion of the graft and the optic disc at the same time as transplantation, in addition to labelled cells situated peripheral to the graft, retrograde labelling with horseradish peroxidase demonstrated a population of labelled neurons located between the graft and the optic disc which was not observed in animals without the additional lesion. Since the axons of this population of cells would have to turn around away from their normal course towards the optic disc and travel for about 1.5 mm in order to grow into the graft, it suggests that the peripheral nerve graft might play an active role in attracting and/or guiding damaged ganglion cell axons to grow into it.
Recent experiments using adult rodents have demonstrated that d a m a g e d retinal ganglion cell axons can regenerate for a long distance into a segment of autologous peripheral nerve graft inserted into the eye of rats 9 or hamsters t~. Limited growth of such axons has also been observed when the graft was placed into the optic tract of hamsters 1°. All these experiments agree with the suggestion that the peripheral nervous system (PNS) environment is favourable for supporting extensive regrowth of axons in the adult mammalian CNS 1. H o w e v e r , it is not clear how the d a m a g e d ganglion cell axons grew into the graft in the first place. It is possible to suggest that the graft might play an active role in attracting and/or guiding the regenerating axons to grow into it. Or the graft might play a passive role in the sense that it intercepts cut ganglion cell axons located peripheral to the graft which are trying to grow towards the direction of the optic disc. It is, however, difficult to differentiate between these two possibilities in the previous experiments 9'n. In o r d e r to investigate this p r o b l e m further, we have studied the response of ganglion cells whose axons would not normally be intercepted by
the graft. The results suggest that the graft might play an active role in attracting and/or guiding d a m a g e d ganglion cell axons to grow into it. Thirteen young adult golden hamsters (Mesocricetus auratus) were used for the experiments of transplantation of a segment of autologous sciatic nerve into the retina. Animals with PNS graft alone. In 3 hamsters, one end of the PNS graft was inserted into the superior temporal q u a d r a n t of the left eye about 2 mm from the limbus (Figs. 1 and 2A). O n e - t w o months later, horseradish peroxidase ( H R P ) was applied to the other end of the graft which was laid over the skull and the retinae were processed for H R P histochemistry 4. The details of these procedures have been described previously 9.
Animals with PNS graft plus an additional lesion in the retina. In 10 hamsters, a segment of sciatic nerve was inserted into the superior t e m p o r a l retina about 2 m m from the limbus as described before 9. In 7 of them an additional lesion was placed, at the same time as the transplantation of the graft, in the retina between the location of the insertion of the graft and
Correspondence: K.-F. So, Department of Anatomy, Faculty of Medicine, University of Hong Kong, Hong Kong. 0006-8993/86/$03.50 O 1986 Elsevier Science Publishers B.V. (Biomedical Division)
169
Fig. 1. Photograph of an eye from a hamster showing the optic nerve (ON) and the graft. The star (*) denotes the location where the graft is inserted into the retina.
the optic disc with the tip of a 22-gauge needle (Fig. 2C). One to two months later, H R P retrograde labelling experiments were conducted in 5 of these animals and the retinae were processed for H R P histochemical reaction as described above. In two animals, double fluorescent dyes labelling experiments were performed. The cytoplasmic label True Blue (2%) was applied to the cut end of the graft and 6 days later the cell nucleus label Nuclear Yellow (2%) was applied to the surgically exposed and transected contralateral (n = 1) or ipsilateral (n = 1) optic tract s (Fig. 3A). Two days later, the animals were perfused with cold normal saline. Both retinae were removed, fixed in 10% formol-saline solution for one hour, fiatmounted, dried, coated with DPX, coverslipped and examined with a Zeiss fluorescent microscope. In the remaining 3 animals with a graft in the retina, an additional lesion was placed in the ventral retina ca. 3 mm away from the graft insertion (Fig. 4). One to two months later, H R P was applied to the graft and the retinae were processed for H R P reaction. The transplanted PNS nerve was found to be well attached to the eye (Fig. 1) and subcutaneous tissues overlying the skull after a postgrafting period of 1-2 months. Animals with peripheral nerve transplantation alone. Retrograde H R P labelling experiments were carried out in animals with a segment of PNS nerve inserted into the left retina. H R P applied to the graft
(Fig. 2A) demonstrated, similar to the results obtained from the rat 9, a population of labelled ganglion cells of different sizes situated peripheral to the insertion of the graft in the fiat-mounted retina (Fig. 2B). No cell was observed in the area between the graft and the optic disc and only a few cells were found on the temporal side of the graft (Figs. 2B and 4). Since it is known that the axons would have to be damaged before they would grow into the graft 9'11, it seems to suggest that, similar to the results observed in the cat retina 5, very few ganglion cell axons follow a curved course to reach the optic disc. No labelled neuron was detected in the control, right, retinae. Animals with peripheral nerve transplantation plus an additional lesion. In 7 animals, an additional damage was placed between the insertion of the graft and the optic disc at the same time of transplantation. When H R P was applied to the graft of 5 of these animals (Fig. 2C), in addition to labelled cells situated peripheral to the graft (between 11 and 276 cells, mean + S.D. = 100 + 103), a population of labelled neurons was observed (between 3 and 140 cells, mean + S.D. = 39 + 58) between the graft and the optic disc (Fig. 2D) which was not observed in animals without the additional lesion. A double fluorescent dyes experiment was conducted in the remaining two animals to investigate the origins of the fibers in the graft from the population of cells located between the graft and site of the additional lesion. The experiment was designed to find out whether they were regenerative outgrowth from axotomized neurons or collateral sprouts from uninjured neurons which still retain their axons in the optic tract. If the second possibility is valid, double labelled neurons should be observed after application of True Blue to the graft and Nuclear Yellow to the contralateral (cell B in Fig. 3A) or ipsilateral optic tract. No such cell was observed in the two animals studied suggesting that the axons in the graft originated from neurons whose axons have been damaged. This conclusion is similar to that obtained for the population of regenerating ganglion cells situated peripheral to the graft 9A1. In 3 additional animals, an additional lesion was placed in the ventral retina, ca. 3 mm from the insertion of the graft. After a postgrafting period of 1-2 months, H R P was applied to the graft and labelled neurons were observed peripheral to the graft but no-
170
B
A graft
HRP
T
eye
OC
contra OT
g
I1 HRP
Fig. 2. A: schematic diagram illustrating retrograde HRP labelling experiment in animals with a segment of sciatic nerve transplanted to the left eye. ON, optic nerve; OC, optic chiasm; contra OT, contralateral optic tract. B: diagram of retinal whole mount of a hamster from an experiment (HRR-4, 44 days postgrafting survival time) illustrated in A. Black dots are HRP labelled neurons; g, graft; U, upper retina; T, temporal retinal; x, optic disc. Scale bar, 1 mm. C: schematic diagram illustrating retrograde HRP labelling experiment in animal with PNS graft plus an additional damage (black bar) in the retina. D: diagram of a retinal whole mount of a hamster from an experiment (DOHRR-5, 30 days post grafting survival time) illustrated in C. d, additional damage in the retina. Other abbreviations, same as in B. Note that HRP labelled neurons (black dots) are also found in areas of the retina between the graft and optic disc. Scale bar, 1 mm.
where else (Fig. 4). In all of the above experiments, no labelled n e u r o n was observed in the control, right, retinae. The results from the present experiments in adult hamsters with a graft and an additional lesion in the retina demonstrated that some cut axons which are not in direct apposition with the graft can also regrow into the graft. This was shown by the labelling of a population of n e u r o n s located between the insertion
of the graft and the site of the additional lesion in the retina. Since the axons of this population of cells would have to turn around away from its normal course towards the optic disc, and travel for ca. 1.5 mm from the site of the additional lesion in order to grow into the graft, it suggests that the grafted peripheral nerve might play an active role in attracting and/or guiding damaged ganglion cell axons to grow into it.
171
B
u
A TB
T
Fig. 3. A: schematic diagram illustrating the double fluorescent dyes experiment in animals with PNS graft plus an additional damage (black bar) in the retina. True Blue (TB) was applied to the graft and Nuclear Yellow (NY) to the optic tract in order to trace, in areas between the graft and optic disc, any neuron which has projections in the graft and optic tract (cell B). Cell A is a cell which sends an axon into the graft alone. Cell C is an uninjured neuron which retains its axon in the optic tract. B: diagram of a retinal whole mount of a hamster from an experiment (DOHRR-9, 55 days post grafting survival time) illustrated in A. True Blue-labelled cells (black dots) were located in a fan shaped area peripheral and central to the graft. Nuclear Yellow labelled cells (not illustrated) were located in the remaining part of the retina. Abbreviations as in Fig. 2. Scale bar, 1 mm.
After discrete damage in the retina of adult mouse with no grafting of peripheral nerve, the damaged ax-
ons have b e e n shown to regrow within the retina for a distance of only up to 500 p m 2 or ca. 450 p m 3. But with the addition of the peripheral nerve in the hamster, the damaged ganglion cell axons seem to be able to regrow for a much longer distance within the retina in order to grow into the graft. The mechanisms underlying the attracting and/or
T
Fig. 4. Diagram of a retinal whole mount from an animal (FDHRR-2, 38 days post grafting survival time) with a PNS graft and an additional damage in the ventral retina. Abbreviations as in Fig. 2. Scale bar, 1 mm.
guiding role of the grafted peripheral nerve are not clear at the m o m e n t . The PNS nerve could secrete a diffusable chemical into the e n v i r o n m e n t to attract the damaged axons. For example, denervated peripheral nerves have been shown to secrete nerve growth factor-like substance 6 and nerve growth associated protein 7. O n the other hand, it is also possible that Schwann cells and/or other types of cells might migrate from the graft into the retina thus providing a substrate to guide the axons to grow into the graft. For example, the migration of Schwann cells, fibroblasts and endothelial cells from cut ends of peripheral nerve into a silicone chamber have b e e n observed to precede that of regenerating axons 12. Both of these mechanisms are, however, not mutually exclusive of each other. But if they operate in the hamster, they seem to be effective in attracting and/or guiding
172 d a m a g e d axons only within a certain r a n g e of dis-
This investigation was s u p p o r t e d by the C r o u c h e r
tance, b e c a u s e axons which h a v e b e e n d a m a g e d ca. 3
F o u n d a t i o n of H o n g K o n g . W e t h a n k Mr. K . C . L a u
m m away f r o m the graft could n o t be s h o w n to g r o w
for technical assistance and Miss S. C h u for typing
into the graft.
the manuscript.
1 Aguayo, A.J., Axonal regeneration from injured neurons in the adult mammalian central nervous system. In C.W. Cotman (Ed.), Synaptic Plasticity, The Guilford Press, New York, 1985, pp. 457-484. 2 Goldberg, S. and Frank, B., Will central nervous systems in the adult mammal regenerate after bypassing a lesion? A study in the mouse and chick visual systems, Exp. Neurol., 70 (1980) 675-689. 3 McConnell, P. and Berry, M., Regeneration of ganglion cell axons in the adult mouse retina, Brain Research, 241 (1982) 362-365. 4 Mesulam, M.-M., Tetramethyl benzidine for horseradish peroxidase neurohistochemistry: a non-carcinogenic blue reaction-product with superior sensitivity for visualizing neural afferents and efferents, J. Histochem. Cytochem., 26 (1978) 106-117. 5 Murakami, D., Sesma, M.A. and Rowe, M.H., Characteristics of nasal and temporal retina in Siamese and normally pigmented cats: ganglion cell composition, axon trajectory and laterality of projection, Brain Behav. Evol., 21 (1982) 67-113. 6 Richardson, P.M. and Ebendal, T., Nerve growth activities
in rat peripheral nerve, Brain Research, 246 (1982) 57-64. 7 Skene, P.J.H. and Shooter, E.M., Denervated sheath cells secrete a new protein after nerve injury, Proc. Natl. Acad. Sci. U.S.A., 80 (1983) 4169-4173. 8 So, K.-F. and Schneider, G.E., Abnormal recrossing retinotectal projections after early lesions in Syrian hamster: age-related effects, Brain Research, 147 (1978) 277-295. 9 So, K.-F. and Aguayo, A.J. Lengthy regrowth of cut axons from ganglion cells after peripheral nerve transplantation into the retina of adult rat, Brain Research, 328 (1985) 349-354. 10 Stevenson, J.A., Growth of optic tract axons in nerve grafts in hamsters, Exp. Neurol., 87 (1985) 446-457. 11 Xiao, Y.-M. and So, K.-F., Ganglion cells regenerating axons into peripheral nerve graft transplanted into the retina of adult hamsters, Hong Kong Soc. Neurosci. Abstr., 7 (1985) 13. 12 Williams, L.R., Longo, F.M., Powell, H.C., Lundborg, G. and Varon, S., Spatial-temporal progress of peripheral nerve regeneration within a silicone chamber: parameters for a bioassay, J. Comp. Neurol., 218 (1983) 460-470.