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A quantitative electron microscopic analysis of the infraorbital nerve in the newborn rat
BrainResearch, 322(1984)369-373 Elsevier
369
BRE 20485
A quantitative electron microscopic analysis of the infraorbital nerve in the newborn rat WILLIAM E. RENEHAN and ROBERT W. RHOADES Department of Anatomy, University of Medicine and Dentistry of New Jersey, School of Osteopathic Medicine and Rutgers Medical School, Piscataway, NJ 08854 (U.S.A.) (Accepted July 10th, 1984) Key words: rat - - infraorbital nerve - - trigeminal system - - development - - electron microscopy
The infraorbital nerve (n = 3) was examined in newborn rats using electron microscopic techniques. Counts of the entire nerve revealed an average of 42,051 (S.D. = 2083) unmyelinated and 168 (S.D. = 47) myelinated fibers. The unmyelinated axons averaged 0.46/~m (S.D. = 0.16) in diameter while the myelinated fibers averaged 1.71 #m (S.D. = 0.17). The infraorbital (IO) nerve, a branch of the maxillary division of the trigeminal (V) nerve, provides the sole afferent innervation of the rodent mystacial vibrissae. It is well known that there is an isomorphic relationship between the facial pattern of these whiskers and their representations in the central nervous system. This highly ordered topography provides an excellent opportunity to study development and plasticity. Numerous studies have shown that vibrissae follicle lesions2.3, 9-11 or transection of the IO nerve 7A2,172° on or soon after birth results in altered topography of the Vth nerve in brainstem, thalamus and cortex. Surprisingly, little is known of the composition of the rat IO nerve at this stage in development. Jacquin et al. have recently shown that the adult IO nerve is composed of 19,740 (S.D. = 2054) myelinated and 13,319 (S.D. = 1159) unmyelinated axonsS. In the present study, we have used electron microscopic methods to delineate the organization and composition of the IO nerve on the day of birth. Rats were deeply anesthetized with sodium pentobarbital (Nembutal) within 6 h of birth and perfused transcardiaily with 0.9% saline containing 0.4% heparin and 0.4% xylocaine, followed by a 2% glutaraldehyde-2% paraformaldehyde fixative in sodium phosphate buffer. Both IO nerves were removed 1 h following the perfusion, rinsed, and placed in a 1.0%
osmium tetroxide post-fixative containing 1.5% potassium ferricyanide15 for 2 h. Following dehydration in graded alcohols, the tissue was embedded in epoxy resin (EM bed-812, Electron Microscopy Sciences) and thick (0.5/~m) and thin (60 nm) sections were taken rostral to the level of the anterior superior alveolar foramen. Thick sections were collected on microscope slides and stained by the technique of Laczko and Levai 13. Thin sections were placed on formvar-coated copper slot grids, stained with lead citrate and aqueous uranyl acetate, and viewed with either a Philips 300 or Philips 400 transmission electron microscope. Three IO nerves were photographed at 1250 x and printed at a final magnification of 8000 x. A complete montage of each nerve was constructed from these prints, and all myelinated and unmyelinated axons were counted. Portions of the nerves (see Fig. 2 A - C ) were photographed at 2750 or 6000 x and enlarged to 16,500 and 36,000 x respectively. These higher magnification prints were used to test the accuracy of the fiber counts in the lower magnification images and for axon area measurements. A n Apple II plus computer with graphics tablet was used to calculate individual axon areas and to convert these values to average diameters (2 x V'-~-~). Each of the IO nerves (Figs. 1A and 2 A - C ) had a triangular shape and was composed of 8 to 15 fas-
Correspondence: W. E, Renehan, Department of Anatomy, UMDNJ-SOM, P.O. Box 55, Piscataway, NJ 08854, U.S.A. 0006-8993/84/$03.00 © 1984 Elsevier Science Publishers B.V.
370
A
Fig. 1. A: thick (0.5/~m) epoxy section of the infraorbital nerve of a newborn rat, rostral to the level of the anterior superior alveolar foramen. Calibration bar equals 50/~m. B: this high magnification electron micrograph illustrates a number of unmyelinated axons and their associated Schwann cells. The membrane-delimited axons were easily distinguished from the surrounding Schwann cell cytoplasm. Calibration bar equals 1.0 ktm. C: most of the myelinated axons were distributed fairly evenly throughout the nerve. As demonstrated in this micrograph, however, the myelinated axons were occasionally encountered in groups of 10-20. Calibration bar equals 1.0/~m.
371
D 4O( A
30~ 20O
.
e
.
'
1.2
1.,.,-
1.4
E
40 C
20 i / 110
1.2
1A Axon
. . . . . 1'.6 1.8 Diameter
2D
L_, 2.2
2.4
(p)
Fig. 2. A-C: line drawings of the 3 IO nerves used in this study, illustrating the size and number of the major nerve fascicles. The boxes indicate the areas chosen for higher magnification analysis of axon diameter and number. D: histogram shows the size distribution for 1580unmyelinated axons in 3 nerves. The mean diameter was 0.46/~m (S.D. = 0.16). E: illustrates the diameter distribution of 250 myelinated axons in 3 nerves. The average diameter was 1.71/tm (S.D. = 0.17). t cicles. Myelinated axons were quite sparse and tended to be distributed fairly evenly throughout the cross section of the nerve. Occasionally, however, they were found in small groups ranging between 10 and 20 in number (Fig. 1C). The vast majority of the axons in the nerve were unmyelinated and readily distinguishable from the surrounding Schwann cell cytoplasm by their lesser density and the presence of a delimiting unit membrane (Fig. 1B). Frequently, a number of unmyelinated axons occupied a single trough of the Schwann cell cytoplasm, The IO nerve in the newborn rat contained an average of 42,051 (S.D. = 2083) unmyelinated and 168 (S.D. = 47) myelinated fibers. The average diameter of the unmyelinated fibers was 0.46/~m (S.D. = 0.16) and the fiber diameter distribution (Fig. 2D) was unimodal. The average diameter for the myelinated axons (Fig. 2E) was 1.7 ~m (S.D. = 0.17). Again, the distribution was unimodal,
The present findings, when compared with those of Jacquin et al. 8 (which report an adult IO complement of 33,000 axons) demonstrate that at least the adult complement of IO nerve fibers have extended significantly toward and probably reached the periphery by the time the rat is born. In this regard, our data agree with those which Davies and Lumsden 4 recently provided for the maxillary nerve of the mouse. Our findings are also consistent with the fact that the vast majority of V ganglion neurons undergo their final divisions before embryonic day (E) 13 and that this neurogenesis is complete by El56. Erzurumlu and Killackey's 5 observation that numerous maxillary nerve fibers extend toward and reach the periphery during embryonic life also supports the conclusion that the adult complement of IO axons have extended well out of the ganglion by the time the rat (and probably also the mouse, see refs. 18 and 19 for additional data) is born. Importantly, all of these
372 data, especially those from the electron microscopic studies, indicate that the IO reinnervation of the
mouse is on E l l lB. Secondly, silver staining in
whiskerpad following neonatal section of this nerve is probably n o t the result of late growing fibers which escape transection. Our data also suggest the possibility that the IO
riphery by E l l , while, in rat, this does not occur until El45. Finally, 7% of the axons in the maxillary nerve
nerve in newborn rats may contain as many as 10,000 more fibers in the n e o n a t e than in the adult animal. These data are consistent with those which have been obtained for other peripheralH6 and central t4~5,21 nerves in mammals. Davies and Lumsden have similarly shown that the maxillary nerve of fetal mice contains excess axons, but that adult fiber n u m b e r s are reached by the end of embryonic development 4. While the sample size in their study is also small (only one nerve and ganglion were counted at E l 9 and these data were compared with ganglion cell counts made at postnatal day 4) there are other data which suggest that the trigeminal system in the mouse may mature slightly earlier than that in the rat. First, peak neurogenesis in the rat V ganglion occurs between E12 and E136 while that in
1 Aguayo, A. J., Terry, L. C. and Bray, G. M., Spontaneous loss of axons in sympathetic unmyelinated nerve fibers of the rat during development, Brain Research, 54 (1973) 360-364. 2 Belford, G. R. and Killackey, H. P., The development of vibrissae representation in subcortical trigeminal centers of the neonatal rat, J. comp. Neurol., 188 (1979) 63-74. 3 Beldford, G. R. and Killackey, H. P., The sensitive period in the development of the trigeminal system of the neonatal rat, J. comp. Neurol., 193 (1980) 335-350. 4 Davies, A. and Lumsden, A., Relation of target encounter and neuronal death to nerve growth factor responsiveness in the developing mouse trigeminal ganglion, J. comp. Neurol., 223 (1984) 124-137. 5 Erzurumlu, R. S. and Killackey, H. P., Development of order in the rat trigeminal system, J. cornp. Neurol., 213 (1983) 365-380. 6 Forbes, D. J. and Welt, C., Neurogenesis in the trigeminal ganglion of the albino rat: a quantitative autoradiographic study, J. comp. Neurol., 199 (1981) 133-147. 7 Jacquin, M. F. and Rhoades, R. W., Central projections of the normal and 'regenerate' infraorbital nerve in adult rats subjected to neonatal unilateral infraorbital lesions: a transganglionic horseradish peroxidase study, Brain Research, 269 (1983) 137-144. 8 Jacquin, M. F., Hess, A., Yang, G., Adamo, P., Math, M., Brown, A. and Rhoades, R. W., Organization of the infraorbital nerve in rat: a quantitative electron microscopic study, Brain Research, 290 (1984) 131-135. 9 Jeanmonod, D., Rice, F. L. and Van der Loos, H., Mouse somatosensory cortex: alterations in the barrelfield follow-
mouse 4 reveals that maxillary axons reach the pe-
are myelinated in n e w b o r n mice 4 while this is true of only 0.4% of the fibers in newborn rats. Our findings for the rat further suggest that the pattern representing the whiskers in the brainstem becomes apparent3 at an age when there are many extra fibers in the IO nerve and when the morphology (i.e. size and degree of myelination) of those fibers is quite immature. Supported by DE06528, EY04170, EY03546, The March of Dimes and the U M D N J Foundation. W . E . R . is the recipient of N R S A NS07444-01. Thanks to A n n Marie Szczepanik for technical assistance and to E l e a n o r Kells and Patti Vendula for typing the manuscript. We are grateful to Dr. Bryce Munger for use of the Philips 400 transmission electron microscope at the M.S. Hershey Medical Center of the Pennsylvania State University.
ing receptor injury at different early postnatal ages, Neuroscience, 6 (1981) 1503-1535.
10 Killackey, H. P., Belford, G., Ryugo, R. and Ryugo, D., Anomalous organization of thalamocortical projections consequent to vibrissae removal in newborn rat and mouse, Brain Research, 183 (1980) 205-210. 11 Killackey, H. P. and Belford, G. R., Central correlates of peripheral pattern alterations in the trigeminal system of the rat, Brain Research, 183 (1980) 205-210. 12 Killackey, H. P. and Shinder, A., Central correlates of peripheral pattern alterations in the trigeminal system of the rat. II. The effect of nerve section, Develop. Brain Res., 1 (1981) 121-126. 13 Laczko, J. and Levai, G., A simple differential staining method for semi-thin sections of ossifying cartilage and bone tissue embedded in epoxy resin, Mikroskopie, 31 (1975) 1-14. 14 Lam, K., Jervie Sefton, A. and Bennett, M. R., Loss of axons from the optic nerve of the rat during early postnatal development, Develop. Brain Res., 3 (1982) 487-491. 15 Langford, L.A. and Coggeshall, R. E., The use of potassium ferricyanide in neural fixation, Anat. Rec., 197 (1980) 297-303. 16 Rakic, P. and Riley, K. P., Overproduction and elimination of retinal axons in the fetal rhesus monkey, Science, 219 (1983) 1441-1444. 17 Reier, P. J. and Hughes, A., Evidence for spontaneous axon degeneration during peripheral nerve maturation, Amer..J. Anat., 135 (1972) 147-152. 18 Rhoades, R. W., Fiore, J. M., Math, M. F. and Jacquin, M. F., Reorganization of trigeminal primary afferents follow-
373 ing neonatal infraorbital nerve section in hamster, Develop. Brain Res., 7 (1983) 337-342. 19 Taber Pierce, E., Histogenesis of the sensory nuclei of the trigeminal nerve in the mouse. An autoradiographic study, Anat. Rec., 166 (1970) 388. 20 Van Exan, R. and Hardy, M., A spatial relationship between innervation and the early differentiation of vibrissa follicles in the embryonic mouse, J. Anat., 131 (1980) 643-656. 21 Waite, P.M. and Cragg, B. G., The peripheral and central changes resulting from cutting or crushing the afferent
nerve supply to the whiskers, Proc. roy. Soc. B, 214 (1982) 191-211. 22 Williams, R. W., Bastiani, M. J. and Chalupa, L. M., Loss of axons in the cat optic nerve following fetal unilateral enucleation: an electron microscopic analysis, J. Neuroscience, 3 (1983) 133-144. 23 Woolsey, T. A. and Van der Loos, H., The structural organization of layer IV in the somatosensory region (SI) of mouse cerebral cortex: the description of a cortical field composed of discrete cytoarchitectonic units, Brain Research, 17 (1970) 205-242.
369
BRE 20485
A quantitative electron microscopic analysis of the infraorbital nerve in the newborn rat WILLIAM E. RENEHAN and ROBERT W. RHOADES Department of Anatomy, University of Medicine and Dentistry of New Jersey, School of Osteopathic Medicine and Rutgers Medical School, Piscataway, NJ 08854 (U.S.A.) (Accepted July 10th, 1984) Key words: rat - - infraorbital nerve - - trigeminal system - - development - - electron microscopy
The infraorbital nerve (n = 3) was examined in newborn rats using electron microscopic techniques. Counts of the entire nerve revealed an average of 42,051 (S.D. = 2083) unmyelinated and 168 (S.D. = 47) myelinated fibers. The unmyelinated axons averaged 0.46/~m (S.D. = 0.16) in diameter while the myelinated fibers averaged 1.71 #m (S.D. = 0.17). The infraorbital (IO) nerve, a branch of the maxillary division of the trigeminal (V) nerve, provides the sole afferent innervation of the rodent mystacial vibrissae. It is well known that there is an isomorphic relationship between the facial pattern of these whiskers and their representations in the central nervous system. This highly ordered topography provides an excellent opportunity to study development and plasticity. Numerous studies have shown that vibrissae follicle lesions2.3, 9-11 or transection of the IO nerve 7A2,172° on or soon after birth results in altered topography of the Vth nerve in brainstem, thalamus and cortex. Surprisingly, little is known of the composition of the rat IO nerve at this stage in development. Jacquin et al. have recently shown that the adult IO nerve is composed of 19,740 (S.D. = 2054) myelinated and 13,319 (S.D. = 1159) unmyelinated axonsS. In the present study, we have used electron microscopic methods to delineate the organization and composition of the IO nerve on the day of birth. Rats were deeply anesthetized with sodium pentobarbital (Nembutal) within 6 h of birth and perfused transcardiaily with 0.9% saline containing 0.4% heparin and 0.4% xylocaine, followed by a 2% glutaraldehyde-2% paraformaldehyde fixative in sodium phosphate buffer. Both IO nerves were removed 1 h following the perfusion, rinsed, and placed in a 1.0%
osmium tetroxide post-fixative containing 1.5% potassium ferricyanide15 for 2 h. Following dehydration in graded alcohols, the tissue was embedded in epoxy resin (EM bed-812, Electron Microscopy Sciences) and thick (0.5/~m) and thin (60 nm) sections were taken rostral to the level of the anterior superior alveolar foramen. Thick sections were collected on microscope slides and stained by the technique of Laczko and Levai 13. Thin sections were placed on formvar-coated copper slot grids, stained with lead citrate and aqueous uranyl acetate, and viewed with either a Philips 300 or Philips 400 transmission electron microscope. Three IO nerves were photographed at 1250 x and printed at a final magnification of 8000 x. A complete montage of each nerve was constructed from these prints, and all myelinated and unmyelinated axons were counted. Portions of the nerves (see Fig. 2 A - C ) were photographed at 2750 or 6000 x and enlarged to 16,500 and 36,000 x respectively. These higher magnification prints were used to test the accuracy of the fiber counts in the lower magnification images and for axon area measurements. A n Apple II plus computer with graphics tablet was used to calculate individual axon areas and to convert these values to average diameters (2 x V'-~-~). Each of the IO nerves (Figs. 1A and 2 A - C ) had a triangular shape and was composed of 8 to 15 fas-
Correspondence: W. E, Renehan, Department of Anatomy, UMDNJ-SOM, P.O. Box 55, Piscataway, NJ 08854, U.S.A. 0006-8993/84/$03.00 © 1984 Elsevier Science Publishers B.V.
370
A
Fig. 1. A: thick (0.5/~m) epoxy section of the infraorbital nerve of a newborn rat, rostral to the level of the anterior superior alveolar foramen. Calibration bar equals 50/~m. B: this high magnification electron micrograph illustrates a number of unmyelinated axons and their associated Schwann cells. The membrane-delimited axons were easily distinguished from the surrounding Schwann cell cytoplasm. Calibration bar equals 1.0 ktm. C: most of the myelinated axons were distributed fairly evenly throughout the nerve. As demonstrated in this micrograph, however, the myelinated axons were occasionally encountered in groups of 10-20. Calibration bar equals 1.0/~m.
371
D 4O( A
30~ 20O
.
e
.
'
1.2
1.,.,-
1.4
E
40 C
20 i / 110
1.2
1A Axon
. . . . . 1'.6 1.8 Diameter
2D
L_, 2.2
2.4
(p)
Fig. 2. A-C: line drawings of the 3 IO nerves used in this study, illustrating the size and number of the major nerve fascicles. The boxes indicate the areas chosen for higher magnification analysis of axon diameter and number. D: histogram shows the size distribution for 1580unmyelinated axons in 3 nerves. The mean diameter was 0.46/~m (S.D. = 0.16). E: illustrates the diameter distribution of 250 myelinated axons in 3 nerves. The average diameter was 1.71/tm (S.D. = 0.17). t cicles. Myelinated axons were quite sparse and tended to be distributed fairly evenly throughout the cross section of the nerve. Occasionally, however, they were found in small groups ranging between 10 and 20 in number (Fig. 1C). The vast majority of the axons in the nerve were unmyelinated and readily distinguishable from the surrounding Schwann cell cytoplasm by their lesser density and the presence of a delimiting unit membrane (Fig. 1B). Frequently, a number of unmyelinated axons occupied a single trough of the Schwann cell cytoplasm, The IO nerve in the newborn rat contained an average of 42,051 (S.D. = 2083) unmyelinated and 168 (S.D. = 47) myelinated fibers. The average diameter of the unmyelinated fibers was 0.46/~m (S.D. = 0.16) and the fiber diameter distribution (Fig. 2D) was unimodal. The average diameter for the myelinated axons (Fig. 2E) was 1.7 ~m (S.D. = 0.17). Again, the distribution was unimodal,
The present findings, when compared with those of Jacquin et al. 8 (which report an adult IO complement of 33,000 axons) demonstrate that at least the adult complement of IO nerve fibers have extended significantly toward and probably reached the periphery by the time the rat is born. In this regard, our data agree with those which Davies and Lumsden 4 recently provided for the maxillary nerve of the mouse. Our findings are also consistent with the fact that the vast majority of V ganglion neurons undergo their final divisions before embryonic day (E) 13 and that this neurogenesis is complete by El56. Erzurumlu and Killackey's 5 observation that numerous maxillary nerve fibers extend toward and reach the periphery during embryonic life also supports the conclusion that the adult complement of IO axons have extended well out of the ganglion by the time the rat (and probably also the mouse, see refs. 18 and 19 for additional data) is born. Importantly, all of these
372 data, especially those from the electron microscopic studies, indicate that the IO reinnervation of the
mouse is on E l l lB. Secondly, silver staining in
whiskerpad following neonatal section of this nerve is probably n o t the result of late growing fibers which escape transection. Our data also suggest the possibility that the IO
riphery by E l l , while, in rat, this does not occur until El45. Finally, 7% of the axons in the maxillary nerve
nerve in newborn rats may contain as many as 10,000 more fibers in the n e o n a t e than in the adult animal. These data are consistent with those which have been obtained for other peripheralH6 and central t4~5,21 nerves in mammals. Davies and Lumsden have similarly shown that the maxillary nerve of fetal mice contains excess axons, but that adult fiber n u m b e r s are reached by the end of embryonic development 4. While the sample size in their study is also small (only one nerve and ganglion were counted at E l 9 and these data were compared with ganglion cell counts made at postnatal day 4) there are other data which suggest that the trigeminal system in the mouse may mature slightly earlier than that in the rat. First, peak neurogenesis in the rat V ganglion occurs between E12 and E136 while that in
1 Aguayo, A. J., Terry, L. C. and Bray, G. M., Spontaneous loss of axons in sympathetic unmyelinated nerve fibers of the rat during development, Brain Research, 54 (1973) 360-364. 2 Belford, G. R. and Killackey, H. P., The development of vibrissae representation in subcortical trigeminal centers of the neonatal rat, J. comp. Neurol., 188 (1979) 63-74. 3 Beldford, G. R. and Killackey, H. P., The sensitive period in the development of the trigeminal system of the neonatal rat, J. comp. Neurol., 193 (1980) 335-350. 4 Davies, A. and Lumsden, A., Relation of target encounter and neuronal death to nerve growth factor responsiveness in the developing mouse trigeminal ganglion, J. comp. Neurol., 223 (1984) 124-137. 5 Erzurumlu, R. S. and Killackey, H. P., Development of order in the rat trigeminal system, J. cornp. Neurol., 213 (1983) 365-380. 6 Forbes, D. J. and Welt, C., Neurogenesis in the trigeminal ganglion of the albino rat: a quantitative autoradiographic study, J. comp. Neurol., 199 (1981) 133-147. 7 Jacquin, M. F. and Rhoades, R. W., Central projections of the normal and 'regenerate' infraorbital nerve in adult rats subjected to neonatal unilateral infraorbital lesions: a transganglionic horseradish peroxidase study, Brain Research, 269 (1983) 137-144. 8 Jacquin, M. F., Hess, A., Yang, G., Adamo, P., Math, M., Brown, A. and Rhoades, R. W., Organization of the infraorbital nerve in rat: a quantitative electron microscopic study, Brain Research, 290 (1984) 131-135. 9 Jeanmonod, D., Rice, F. L. and Van der Loos, H., Mouse somatosensory cortex: alterations in the barrelfield follow-
mouse 4 reveals that maxillary axons reach the pe-
are myelinated in n e w b o r n mice 4 while this is true of only 0.4% of the fibers in newborn rats. Our findings for the rat further suggest that the pattern representing the whiskers in the brainstem becomes apparent3 at an age when there are many extra fibers in the IO nerve and when the morphology (i.e. size and degree of myelination) of those fibers is quite immature. Supported by DE06528, EY04170, EY03546, The March of Dimes and the U M D N J Foundation. W . E . R . is the recipient of N R S A NS07444-01. Thanks to A n n Marie Szczepanik for technical assistance and to E l e a n o r Kells and Patti Vendula for typing the manuscript. We are grateful to Dr. Bryce Munger for use of the Philips 400 transmission electron microscope at the M.S. Hershey Medical Center of the Pennsylvania State University.
ing receptor injury at different early postnatal ages, Neuroscience, 6 (1981) 1503-1535.
10 Killackey, H. P., Belford, G., Ryugo, R. and Ryugo, D., Anomalous organization of thalamocortical projections consequent to vibrissae removal in newborn rat and mouse, Brain Research, 183 (1980) 205-210. 11 Killackey, H. P. and Belford, G. R., Central correlates of peripheral pattern alterations in the trigeminal system of the rat, Brain Research, 183 (1980) 205-210. 12 Killackey, H. P. and Shinder, A., Central correlates of peripheral pattern alterations in the trigeminal system of the rat. II. The effect of nerve section, Develop. Brain Res., 1 (1981) 121-126. 13 Laczko, J. and Levai, G., A simple differential staining method for semi-thin sections of ossifying cartilage and bone tissue embedded in epoxy resin, Mikroskopie, 31 (1975) 1-14. 14 Lam, K., Jervie Sefton, A. and Bennett, M. R., Loss of axons from the optic nerve of the rat during early postnatal development, Develop. Brain Res., 3 (1982) 487-491. 15 Langford, L.A. and Coggeshall, R. E., The use of potassium ferricyanide in neural fixation, Anat. Rec., 197 (1980) 297-303. 16 Rakic, P. and Riley, K. P., Overproduction and elimination of retinal axons in the fetal rhesus monkey, Science, 219 (1983) 1441-1444. 17 Reier, P. J. and Hughes, A., Evidence for spontaneous axon degeneration during peripheral nerve maturation, Amer..J. Anat., 135 (1972) 147-152. 18 Rhoades, R. W., Fiore, J. M., Math, M. F. and Jacquin, M. F., Reorganization of trigeminal primary afferents follow-
373 ing neonatal infraorbital nerve section in hamster, Develop. Brain Res., 7 (1983) 337-342. 19 Taber Pierce, E., Histogenesis of the sensory nuclei of the trigeminal nerve in the mouse. An autoradiographic study, Anat. Rec., 166 (1970) 388. 20 Van Exan, R. and Hardy, M., A spatial relationship between innervation and the early differentiation of vibrissa follicles in the embryonic mouse, J. Anat., 131 (1980) 643-656. 21 Waite, P.M. and Cragg, B. G., The peripheral and central changes resulting from cutting or crushing the afferent
nerve supply to the whiskers, Proc. roy. Soc. B, 214 (1982) 191-211. 22 Williams, R. W., Bastiani, M. J. and Chalupa, L. M., Loss of axons in the cat optic nerve following fetal unilateral enucleation: an electron microscopic analysis, J. Neuroscience, 3 (1983) 133-144. 23 Woolsey, T. A. and Van der Loos, H., The structural organization of layer IV in the somatosensory region (SI) of mouse cerebral cortex: the description of a cortical field composed of discrete cytoarchitectonic units, Brain Research, 17 (1970) 205-242.