J. Vet. Med. B 40, 413-422 (1993) 8 1993 Paul Parey Scientific Publishers, Berlin and Hamburg ISSN 0931 - 1793

Lehrstuhl fur Allgemeine Pathologie und Neuropathologiel (Prof. Dr. E . Dahme)

und II. Medizinische Tierklinikz (Prof. Dr. G . Dirksen)

der Tierarztlichen Fakultat der Universitat Munchen

Spinal Dysmyelination in New-Born Brown Swiss x Braunvieh Calves

ANGEFA HAFNER~, E. DAHME~, GABRIELE OBERMAIER~, P. SCHMIDT~ and G. DIRK SEN^

Address of authors: Dr. ANGELA HAFNER, Institut fur Tierpathologie der Universitat Miinchen, Veterinarstr. 13, 80539 Miinchen, Germany

With 7figures

(Received for publication June 22, 1993)

Summary In four new-born Braunvieh calves suffering from connate recumbency and body tremor, a

hitherto not described myelination disorder of the spinal cord was examined. Bilateral symmetric hypo- as well as demyelination in several spinal tracts were the most conspicuous findings, affecting the ascending gracile funiculus, the ascending dorsolateral spinocerebellar tract, and the mainly descending sulcomarginal tract. Deficient myelin production, loss of myelin, consecutive axonal degenerations, and prominent astrogliosis within these tracts were the histological hallmarks of the disease. This possibly inherited primary myelination disorder of the spinal cord differs markedly from known hereditary neurological diseases in Brown Swiss and Braunvieh cattle, respectively, i. e. the weaver-syndrome and the spinal muscular atrophy.

Introduction In the Braunvieh cattle breed, two hereditary diseases of the central nervous system

(CNS) affecting animals upgraded with American Brown Swiss have been described during the last years: the weaver-syndrome (BRAUN et al., 1987; HAFNER et al., 1991), and the spinal muscular atrophy (SMA) (DAHME et al., 1991; DIRKSEN et al., 1992; STOCKER et al., 1992). The bovine weaver-syndrome is observed in animals aged 6 months to 3 years; the main clinical signs consist of progressive hind limbs ataxia and dysmetria. Calves suffering from SMA show rapidly progressing atrophy of skeletal muscles followed by recumbency at the age of three to seven weeks.

Recently, we observed four Braunvieh calves which were already recumbent at birth exhibiting tremor and general slender profiles of skeletal muscles. The question was raised whether these changes represent only a different clinical course of a known disease (particularly SMA) or constitute a new neurological and neuropathological entity. There- fore, the aim of this study was to examine these cases by light microscopy including immunohistochemistry, and electron microscopy, and to compare the results with the lesions occurring in the above mentioned hereditary diseases.

U.S.Copyright Clearance Center Code Statement: 0931 - 1793/93/4006-0413$02.50/0

414 HAFNER, DAHME, OBERMAIER, SCHMIDT and DIRKSEN

Material and Methods Four Brown Swiss x Braunvieh calves (3 females, 1 male) aged 7, 8, 11, and 12 days (calves no. 1

to 4) were examined. The animals were euthanized and necropsied in the Institute of Veterinary Pathology of the University of Munich.

The brain, spinal cord including spinal ganglia, and specimens of skeletal muscle ( M . biceps fenoris, M . triceps bruchii, M . zntercostulis, and diaphragm) were fixed by immersion in 7 % neutral buffered formalin, and routinely embedded in paraffin. Four coronal brain sections and five segments of the spinal cord (C3, C8, TI, T4, L4) were stained with hematoxylin and eosin (H & E), cresyl violet, and Weigert’s lithium-hematoxylin for myelin staining. Frozen sections of formalin-fixed spinal segments C4 and T5 were stained with oil red 7B. Paraffin sections of skeletal muscles were stained with H & E and with modified Gomori’s trichrome method.

Immunohistochemical examination of spinal segments (C3, C8, T4, L4) was performed on the formalin-fixed paraffin-embedded specimens applying the avidin-biotin complex (ABC) method (Hsu et al., 1981). A monoclonal antibody against 200kD neurofilament protein (Sigma, Deisenhofen, Germany) (dilution 1 : 2,000) and a polyclonal antiserum against glial fibrillary acidic protein (GFAP) (Dako, Hamburg, Germany) (dilution 1 : 100) served as primary antibodies; 3,3’-diaminobenzidine (DAB) was used as enzyme substrate.

The spinal cord segment C7, the sciatic nerve, and the brachial plexus of the 12 day; old calf (no. 4) were fixed by immersion in 6.25 YO glutaraldehyde/Smensen buffer (pH 7.4) for electron microscopy. The dorsal, lateral, and ventral columns of the white matter, the grey matter of spinal cord, and the peripheral nerves were routinely processed and embedded in Epon@. Screening was performed on one pm semithin sections stained with azure blue. Ultrathin sections stained with uranyl acetate and lead citrate were investigated with an electron microscope Zeiss EM 10.

Three age-matched calves (two Braunvieh, one German Simmental calf) which did not show any clinical and pathological signs of neurological diseases served as controls.

Results Gross pathology

At necropsy no gross pathological alterations were observed except for a moderate systemic reduction of skeletal muscle development.

Light microscopy including immunohistochemistry The main histopathological findings occurred at all levels of the spinal cord and were

confined to the white matter. All examined spinal sections showed similar lesions, however with segmental variations. The most pronounced defects were observed in the cervical and thoracic segments. A bilateral symmetric lack of myelin sheaths in the ascending gracile funiculus, in the ascending dorsal spinocerebellar tract, and to a lesser degree, also in the descending sulcomarginal tract was the most obvious alteration (Fig. 1). Within these areas, myelin staining revealed a reduced amount of myelin. Furthermore, bright red staining in oil red preparations indicated additional myelin breakdown.

Immunohistochemical labelling of 2OOkD neurofilament protein revealed a marked numerical reduction of axonal processes in the affected white matter regions but not a complete absence of nerve fibers. However, this was not a lesion as striking as the severe deficient myelination. Additionally, a few spheroids were observed.

Distinct astrocytic gliosis within the myelin-deficient white matter areas was indi- cated by an intense immunohistochemical reaction with the GFAP-antiserum. Thickening of the lamina gliae limitans was a further conspicuous finding.

All these pathohistological findings were markedly evident in the marginal areas of the affected funiculi of the spinal white matter (Fig. 1).

In contrast to the spinal cord, spinal nerve roots and ganglia did not show any pathological alteration. Examination of coronal brain sections comprising the red nucleus, the cuneate and gracile nuclei, and the cerebral motor cortex also did not reveal light microscopic lesions. Especially myelination of the brain, spinal nerve roots and ganglia seemed to be fully matured in accordance with the age-matched controls.

Spinal Dysmyelination in New-Born Brown Swiss x Braunvieh Calves 415

Fig. 1. a. Spinal segment C3, calf no.2: Bilateral symmetric deficit of myelin within the gracile funiculus (L), the dorsal spinocerebellar tract (+), and the sulcomarginal tract (T). Note the intensely stained dorsal and ventral roots. Myelin stain, x 6. b. Spinal segment C3, control calf: Overall intense

staining of the white matter. Myelin stain, x 6

Skeletal muscle tissue was examined in order to exclude a myogenic origin of the disease. The diaphragm and the intercostal muscles displayed a distinct neurogenic atrophy with secondary cellular infiltration, whereas the limb muscles showed an only moderate atrophy of muscle fibers. Thus, no signs of a primary muscular disorder could be detected.

Fig. 2. Spinal segment C7, transversal section of the dorsal column, calf no. 4: Severely affected gracile funiculus which shows obvious lack of myelin sheaths as compared to the adjacent cuneate funiculus (lower left). Most of the remaining myelin sheaths are thinner than would be expected for the

corresponding axon diameter (arrows). Semithin section, azure blue, x 500

416 HAFNER, DAHME, OBERMAIER, SCHMIDT and DIRKSEN

Fig.3. Spinal segment C7, dorsal column, calf no.4: Only a few hypomyelinated axons (a) are interspersed between abundant astroglial processes packed with intermediate filaments. Some of these processes show electron-lucent, hydropic cytoplasm containing peripherally located filament bundles

(asterisk). Transmission electron micrograph (TEM), x 8,000

Electron microscopy The most prominent ultrastructural feature consisted of a severe though not absolute

lack of myelin and, thus, confirmed the light microscopical and immunohistochemical findings. In all affected areas described above, the majority of remaining axons were devoid of regular myelin sheaths (Fig. 2, 3). Longitudinal sections revealed nerve fibers lacking a continuous envelope. Some internodes were naked (Fig. 5), whereas adjacent segments of the same axons were surrounded by a normal-appearing but disproportionately thin myelin sheath or by splitted and vacuolated myelin lamellae. Only a small part of fibers were ensheathed by apparently normal myelin formation. Several nodes of Ranvier were widened (Fig. 4); the adjacent myelin sheaths often were thinner than would be expected for the particular zonal size. The corresponding paranodes showed relatively normal- appearing inward endings of myelin loops. Also heminodes (with only one myelinated paranode bounded to the nodal region) were observed.

In connection with the myelin sheaths, the cell soma of some oligodendrocytes revealed different necrobiotic stages characterized by vacuolation of the cytoplasm and karyorrhectic figures (Fig. 6). However, the total number of oligodendroglial nuclei seemed not to be markedly reduced. A further ultrastructural feature of numerous oligodendrocytes was the distension of the rough endoplasmic reticulum. Myelin debris in the cytoplasm of oligodendroglial cells was encountered in all examined specimens, while no microglial proliferation and only sporadic mononuclear cells were detectable.

The axoplasm of most remaining nerve fibers, even of the hypo- or unmyelinated ones, appeared normal (Fig. 5). Only a few slightly swollen axons showed accumulation

Spinal Dysmyelination in New-Born Brown Swiss x Braunvieh Calves 41 7

Fig. 4. Spinal segment C7, spinocerebellar tract, calf no. 4: Widened node of Ranvier; the swollen, unmyelinated axon is filled with disoriented filaments and tubules (a). The adjacent hypomyelinated parts of the axon (arrows) appear unchanged. Astrocytic processes predominate in the affected area.

TEM, x 8,000

Fig. 5. Spinal segment C7, spinocerebellar tract, calf no. 4: Longitudinal section of an unmyelinated axon with normal appearing axoplasm (a). Abundant astrocytic processes (asterisks) are in direct contact with the axolemma. TEM, x 19,000. Inset: Spinal segment C7, longitudinal section of the lateral column, calf no. 4: Spinocerebellar tract containing unmyelinated (arrows) and hypomyelinated

(arrowhead) axons. Semithin section, azure blue, x 800

418 HAPNER, DAHME, OBERMAIER, SCHMIDT and DIRKSEN

Fig. 6. Spinal segment C7, ventral column, calf no. 4: Necrosis of an oligodendrocyte with karyor- rhectic figure (arrow) and only a few splitted myelin lamellae enveloping a highly swollen axon

containing disoriented filaments (a). TEM, x 3,000

Fig. 7. Spinal segment C7, ventral column, calf no. 4: ‘Reactive axonal enlargement’ packed with lysosomal bodies, mitochondria, and other organelles, and scantily ensheathed by a few myelin

lamellae. TEM, x 3,000

Spinal Dysmyelination in New-Born Brown Swiss x Braunvieh Calves 419

and disorientation of microtubules and intermediate filaments. These changes occurred mostly in axonal zones lacking myelination, whereas, as a rule, adjacent myelinated axonal parts appeared unchanged. A few highly swollen axons and axonal spheroids, respectively, were packed with organelles, especially mitochondria and membrane-bound, electron- dense lysosomal bodies. These so-called ‘reactive axonal enlargements’ (LAMPERT, 1967) or ‘terminal clubs’ (KAO et al., 1977) were covered by extremely thin myelin sheaths (Fig. 7).

The excessive hypertrophy of astrocytic processes that contain densely packed intermediate filaments was a further conspicuous ultrastructural finding within the affected spinal funiculi (Figs. 3, 4, 5). Nerve fibers were interspersed with abundant astroglial processes, and the astrocytic membranes often were in direct contact with naked axons. In none of the investigated cases, astroglial processes occurred between the axon and its corresponding myelin sheath. The astrogliosis was accompanied with scattered astroglial oedema, characterized by segmental vacuolation and swelling, an electron-lucent cyto- plasm and peripherally grouped bundles of filaments (Fig. 3).

The nerve cell bodies of the spinal cord, especially the motoneurons of the ventral horn, appeared unchanged.

Schwann cells and myelin sheaths, respectively, of nerve roots, sciatic nerve, and brachial plexus did not show any pathological alteration, whereas moderate signs of axonal atrophy were observed. These axonal changes were confined to thick myelinated fibers. Nonmyelinated axons of autonomic nerve bundles were unaltered.

Discussion In the present study a hitherto not described spinal disease in Braunvieh calves was

investigated by light microscopy, including immunohistochemistry, and electron micro- scopy.

A severe bilateral symmetric hypomyelination accompanied with demyelination affecting the dorsal, dorsolateral, and ventral spinal tracts were the most conspicuous features observed in four calves suffering from connate recumbency and body tremor.

In general, hypomyelination can be caused by viral infections acquired during fetal life or by exposure to teratogenic substances as well as by genetic defects. In cattle, fetal infection with the BVD-virus is of great importance leading to growth retardation, abortion of the fetus, and neurological disorders in liveborn calves, respectively. Besides various gross pathologic malformations of the brain, hypomyelination is one of the most common findings (BINKHORST et al., 1983). However, to our knowledge, a distribution pattern of myelination deficit as described in our four calves has not been reported so far. In some experimental BVD-cases, an additional spinal dysmyelination could be demon- strated histochemically and biochemically, but neurohistological findings were not de- scribed in detail (DONE et al., 1980).

Inherited hypomyelinating disorders in cattle are known only in a few nosological entities. One of these, the bovine myelinopathy in Australian calves, predominantly of the Murray Grey breed (RICHARDS and EDWARDS, 1986), shares some aspects with the condition described in this report. In both diseases, changes of the spinal white matter are particularly severe in the marginal areas of the ventral and lateral funiculi. However, the dorsal funiculi are affected to a much lesser degree in Murray Grey as compared to our findings in Braunvieh calves. Furthermore, in contrast to our calves, neuronal chromatoly- sis, though probably secondary, is encountered in several brain stem nuclei and spinal grey matter of the Australian cattle.

Two further bovine hereditary myelination disorders differ markedly from the condition described in this report: in Limousin calves, a primary demyelination disorder occurs in a plaque-like, focal pattern affecting predominantly the cerebellar white matter (HARPER et al., 1990; PALMER et al., 1991). In the progressive ataxia of Charolais cattle, eosinophilic plaques composed of hypertrophied oligodendrocytes and disorganized my-

42 0 HAFNER, DAHME, OBERMAIER, SCHMIDT and DIRKSEN

elin sheaths in certain brain and spinal cord regions are the most prominent lesions (BLAKEMORE et al., 1974; CORDY, 1986; PALMER and BLAKEMORE, 1975).

In dogs, there are more reports than in cattle concerning suspicious or proven inherited hypomyelinating disorders, for example in Dalmatian dogs (GREENE et al., 1977), Chow-Chow dogs (VANDEVELDE et al., 1978,1981), Springer spaniel (DUNCAN et al., 1983; GRIFFITHS et al., 1981), Lurcher pups (MAYHEW et al., 1984), Samoyed pups (CUMMINGS et al., 1986), and Weimaraner dogs (KORNEGAY et al., 1987). The striking features of these particular diseases consist of deficient myelination with numerous either unmyelinated or disproportionately hypomyelinated axons within the CNS, in contrast to regular myelin- ation within the peripheral nervous system. In so far, they are in accordance with the changes observed in our bovine cases, however, with the exception that in the canine disorders hypomyelination occurs in both brain and spinal cord. Spinal myelination defects are most obvious at the margins of the ventral and lateral funiculi in our calves and also in some canine cases (KORNEGAY et al., 1987; MAYHEW et al., 1984; VANDEVELDE et d., 1978), whereas the involvement of the dorsal column seems to be an unique histological feature in our bovine cases.

A common finding in several hypomyelinating disorders is a distinct astrocytosis which was also evident in our study. The authors of most of the relevant reports (CUMMINGS et al., 1986; DENTINGER et al., 1982; KORNEGAY et al., 1987; PALMER et al., 1991; RICHARDS and EDWARDS, 1986; VANDEVELDE et al., 1978, 1981) conclude that astrogliosis is rather a consecutive than a primary event. In contrast, astrocytic hypertro- phy was occasionally suspected as a causative factor in hereditary hypomy elination of jimpy mouse (SKOFP, 1976), but this view seems to be not generally accepted (BILLINGS- GAGLIARDI et al., 1980). Since evidence for the existence of a pathogenetic role of astrocytes in the condition described here, e. g. astroglial sprouting between the axon and the corresponding myelin sheath, is lacking, we also consider the astrocytic proliferation as a secondary phenomenon.

Taking into account the particular neurohistological findings, especially the survival of many hypo- and unmyelinated axons as well as the paucity of axonal degenerations in the affected areas of the white matter, the described spinal disorder in Braunvieh calves should be classified as a primary dysmyelination. Consecutive axonal degeneration was observed in the CNS and - to a lesser degree - also in peripheral nerves, while myelin defects selectively had involved the CNS. This finding supports the hypothesis that the disease is due to an impairment of oligodendrocyte function and maturation. In so far it seems to be in accordance with certain canine myelinating disorders which are presumably based on a retarded or abnormal glial differentiation (CUMMINGS et al., 1986; GREENE et al., 1977; KORNEGAY et al., 1987; VANDEVELDE et al., 1978). A further aspect of dysmyelin- ation in our cases is necrosis of oligodendrocytes and myelin breakdown, which might be indicative of oligodendroglial abiotrophy. Unusual oligodendrocytes with distended rough endoplasmic reticulum and nuclear envelope were encountered in our calves alike in the hypomyelinating disorder of Springer spaniel (DUNCAN et al., 1983), and Samoyed pups (CUMMINGS et al., 1986). In these canine diseases, such morphological abnormalities seem not to precede cellular degeneration; they might be signs of increased protein synthesis, disturbed transport and/or “production of an abnormal secretory product” (DUNCAN et al., 1983). However, because of the degenerative changes seen in oligodendrocytes, we cannot exclude that in our cases those alterations represent a precursor-state of cell death.

In comparative terms, the described neurohistological findings differ markedly from those found in pure- and cross-bred Brown Swiss cattle suffering from known hereditary neurological diseases. In contrast to SMA, which is characterized by degeneration and necrosis of the spinal motoneurons (DIRKSEN et al., 1992; EL-HAMIDI et al., 1989; STOCKER et al., 1992), in the present study, nerve cell bodies are lacking any changes. Additionally, the clinical signs of SMA become apparent at the age of two to three weeks (DIRKSEN et al., 1992; EL-HAMIDI et al., 1989; STOCKER et al., 1992), whereas the “dysmyelination calves” showed recumbency and body tremor already at birth. However, it should not be

Spinal Dysmyelination in New-Born Brown Swiss x Braunvieh Calves 42 1

neglected that quite a number of Brown Swiss-upgraded Red Danish Milkbreed calves, which exhibited SMA specific lesions, were reported to be also recumbent perinatally (HANSEN et al., 1988).

Several pathological features observed in the present study are to a certain degree comparable to those revealed in weaver cattle, especially with regard to the funicular pattern of the changes of the spinal white matter including astrogliosis (EL-HAMIDI et al., 1990; STUART and LEIPOLD, 1985). However, electron microscopy of weaver-syndrome suggests a primary axonal degeneration accompanied with a moderate and probably secondary demyelination. Alteration of the axonal mitochondria, which might be of pathogenetic importance in the weaver-syndrome (HAFNER et al., 1991), were not observed in the calves of this study. Therefore, axonal changes, although present in the latter, cannot be considered as a relevant pathognomonic phenomenon. O n the contrary, the absence of conspicuous primary axonal lesions seems to be a common feature of the described condition, in accordance with several bovine and canine primary myelination disorders (BLAKEMORE et d., 1974; CUMMINGS et al., 1986; GREENE et al., 1977; GRIFFITHS et a]., 1981; KORNEGAY et al., 1987; MAYHEW et al., 1984; PALMER et al., 1991; RICHARDS and EDWARDS, 1986; VANDEVELDE et al., 1978, 1981).

In conclusion, we describe a myelination disorder of spinal cord in Braunvieh calves, which hitherto has not been reported in this breed. Although it shares some morphological features with various hereditary bovine and canine diseases, none of these disorders exhibit congruent lesions. Thus, this peculiar bovine dysmyelination should be considered as a new neuropathological entity. A hereditary pathogenesis is suspected, although an infec- tious or toxic etiology cannot be definitely excluded.

Acknowledgements The authors wish to thank Dr. H. SCHWARZ, Peiting, for referring the first case to us, and Miss

CORINNA BEIER and Miss CLAUDIA HOFMANN for excellent technical assistance.

References 1. BILLINGS-GAGLIARDI, S., L. H. ADCOCK, and M. K. WOLF, 1980: Hypomyelinated mutant

mice: description of jpmsd and comparison with jp and qk on their present genetic backgrounds. Brain Res. 194, 325-338.

2. BINKHORST, G. J., D. L. H. JOURNE~, W. WOUDA, P. J. STRAVER, and J. H. Vos, 1983: Neurologi- cal disorders, virus persistence and hypomyelination in calves due to intrauterine infections with bovine virus diarrhoea virus. I. Clinical symptoms and morphological lesions. Vet. Q 5,

3. BLAKEMORE, W.F., A.C. PALMER, and R.M. BARLOW, 1974: Progressive ataxia of Charolais cattle associated with disordered myelin. Acta Neuropathol. 29, 127- 139.

4. BRAUN, U., F.EHRENSPERGER und V.BRACHER, 1987: Das Weaver-Syndrom beim Rind - Klinische, biochemische und pathologisch-anatomische Untersuchungen bei einem Braunvieh-/ Brown Swiss-Rind mit boviner progressiver, degenerativer Myeloenzephalopathie (BPDME). Tierarztl. Prax. 15, 139-144.

5. CORDY, D. R., 1986: Progressive ataxia of Charolais cattle - An oligodendroglial dysplasia. Vet. Pathol. 23, 78-80.

6. CUMMINGS, J. F., B. A. SUMMERS, A. DELAHUNTA, and C. LAWSON, 1986: Tremors in Samoyed pups with oligodendrocyte deficiencies and hypomyelination. Acta Neuropathol. 71,267-277.

7. DAHME, E., A. HAPNER, and P. SCHMIDT, 1991: Spinal muscular atrophy in German Braunvieh calves - comparative neuropathological evaluation. Neuropathol. Appl. Neurobiol. 17, 51 7.

8. DENTINGER, M. P., K. D. BARRON, and C. K. CSIZA, 1982: Ultrastructure of the central nervous system in a myelin deficient rat. J. Neurocytol. 11, 671-691.

9. DIRKSEN, G., K. DOLL, A. HAFNER, W. HERMANNS und E. DAHME, 1992: Spinale Muskelatrophie (SMA) bei Kalbern aus Brown Swiss x Braunvieh-Kreuzungen. Dtsch. tierarztl. Wschr. 99,

10. DONE, J.T., S.TERLECKI, CRICHARDSON, J.W. HARKNESS, J.J. SANDS, D.P.S. PATTERSON, D.SWEASEY, I.G. SHAW, C.E. WINKLER, and S.J. DUFFELL, 1980: Bovine virus diarrhoea- mucosal disease virus: Pathogenicity for fetal calf following maternal infection. Vet. Rec. 106, 473-479.

145 - 155.

168-175.

422 HAFNER, DAHME, OBERMAIER, SCHMIDT and DIRKSEN

11. DUNCAN, I.D., I. R. GRIFFITHS, and M.MuNz, 1983: “Shaking pups”: a disorder of central my- elination in the spaniel dog. 111. Quantitative aspects of glia and myelin in the spinal cord and optic nerve. Neuropathol. Appl. Neurobiol. 9, 355-368.

12. EL-HAMIDI, M., H. W. LEIPOLD, J. G. E. VESTWEBER, and G. SAPERSTEIN, 1989: Spinal muscular atrophy in Brown Swiss calves. J. Vet. Med. A 36, 731-738.

13. EL-HAMIDI, M., H. W. LEIPOLD, and J. E. COOK, 1990: Ultrastructural changes in Brown Swiss cattle affected with Bovine Progressive Degenerative Myeloencephalopathy (Weaver Syndrome). J. Vet. Med. A 37, 729-736.

14. GREENE, C. E., M. VANDEVELDE, and E. J. HOFP, 1977: Congenital cerebrospinal hypo- myelinogenesis in a pup. JAVMA 171, 534-536.

15. GRIFFITHS, I.R., I.D. DUNCAN, and M.MCCULLOCH, 1981: Shaking pups: a disorder of central myelination in the spaniel dog. 11. Ultrastructural observations on the white matter of the cervical spinal cord. J. Neurocytol. 10, 847-858.

16. HAFNER, A., P. SCHMIDT, E. DAHME, and G. DIRKSEN, 1991: The bovine weaver syndrome. Clin. Neuropathol. 10, 34.

17. HANSEN, K. M., H. V. KROGH, J. ENGEL MOLLER, and F. ELLEBY, 1988: Liggekalve-syndromet hos RDM - En ny arvelig kvaegsygdom (The recumbent calf syndrome in Red Danish Milkbreed - A new hereditary disease). Dansk. Vet. Tidsskr. 77, 128-132.

18. HARPER, P.A. W., W. J. HARTLEY, G.C. FRASER, J. G. BOULTON, and N . R. BROWN, 1990: Multifocal encephalopathy in Limousin calves. Austr. Vet. J. 67, 1 1 1 - 112.

19. Hsu, S.-M., L. RAINE, and H. FANGER, 1981: The use of avidin-biotin-peroxidase complex (ABC) in immunoperoxidase technique - a comparison between ABC and unlabeled (PAP) procedu- res. J. Histochem. Cytochem. 29, 577-580.

20. KAO, C. C., L. W. CHANG, and J. M. B. BLOODWORTH, 1977: Electron microscopic observations of the mechanisms of terminal club formation in transected spinal cord axons. J. Neuropathol. Exp. Neurol. 36, 140-156.

21. KORNEGAY, J. N., M.A. GOODWIN, and L. K. SPYRIDAKIS, 1987: Hypomyelination in Weimara- ner dogs. Acta Neuropathol. 72, 394-401.

22. LAMPERT, P. W., 1967: A comparative electron microscopic study of reactive, degenerating, regenerating, and dystrophic axons. J. Neuropathol. Exp. Neurol. 26, 345-368.

23. MAYHEM, I. G., W. F. BLAKEMORE, A. C. PALMER, and C. J. CLARKE, 1984: Tremor syndrome and hypomyelination in Lurcher pups. J. small Anim. Pract. 25, 551 -559.

24. PALMER, A. C., and W. F. BLAKEMORE, 1975: Progressive ataxia of Charolais cattle. Bovine Pract.

25. PALMER, A.C., P.G.G. JACKSON, and W.F. BLAKEMORE, 1991: A primary demyelinating

26. RICHARDS, R. B., and J. R. EDWARDS, 1986: A progressive spinal myelinopathy in beef cattle. Vet.

27. SKOFF, R.P., 1976: Myelin deficit in the Jimpy mouse may be due to cellular abnormalities in

28. STOCKER, H., P. OSSENT, R. HECKMANN und C. OERTLE, 1992: Spinale Muskelatrophie bei

29. STUART, L. D., and H. W. LEIPOLD, 1985: Lesions in bovine progressive degenerative myeloen-

30. VANDEVELDE, M., K. G. BRAUND, T. L. WALKER, and J. N. KORNEGAY, 1978: Dysmyelination of

31. VANDEVELDE, M., K. G. BRAUND, P. J. LUTTGEN, and R. J. HIGGINS, 1981: Dysmyelination in

10, 84-86.

disorder of young cattle. Neuropathol. Appl. Neurobiol. 17,457-467.

Pathol. 23, 35-41.

astroglia. Nature 264, 560-562.

Braunvieh-Kalbern. Schweiz. Arch. Tierheilk. 134, 97- 104.

cephalopathy (“Weaver”) of Brown Swiss cattle. Vet. Pathol. 22, 13-23.

the central nervous system in the Chow-Chow dog. Acta Neuropathol. 42, 211 -215.

Chow Chow dogs: Further studies in older dogs. Acta Neuropathol. 55, 81-87.