Full text of DOI:10.1080/00480169.2002.36312

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Progressive ataxia and seizures in a Cocker Spaniel:
a new type of neurodegenerative disease with novel
intra-neuronal inclusions
RD Jolly a , I Schraa b & TR Halsey a
a Institute of Veterinary, Animal and Biomedical Sciences, Massey University , Palmerston
North, New Zealand E-mail:
b Paremata Veterinary Hospital , 15 Paremata Crescent, Porirua, New Zealand
Published online: 22 Feb 2011.
To cite this article: RD Jolly , I Schraa & TR Halsey (2002) Progressive ataxia and seizures in a Cocker Spaniel: a new type
of neurodegenerative disease with novel intra-neuronal inclusions, New Zealand Veterinary Journal, 50:5, 203-206, DOI:
10.1080/00480169.2002.36312
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New Zealand Veterinary Journal 50(5), 203-206, 2002
203
Clinical Communication
Progressive ataxia and seizures in a Cocker Spaniel: a new type of
neurodegenerative disease with novel intra-neuronal inclusions
RD Jolly*§, I Schraa† and TR Halsey*
occasions over 11 months. On initial examination at 4 months of
age, generalised motor seizures, head tremor and distinct hindlimb
ataxia were evident. Hepatic encephalopathy was discounted as a
likely cause on the basis of serum biochemistry, and the dog was
placed on 30 mg phenobarbitone twice daily. Despite this, neu-
rological signs became progressively worse in ensuing weeks, to a
stage when the dog had 10–15 episodes of seizure activity per day
that would last up to 15 sec. He also began to snap or bite when
handled. At 8 months of age, medication was increased to 60 mg
of phenobarbitone twice daily and 400 mg of potassium bromide
once daily in an attempt to control the tremors and seizures.
Abstract
AIMS: To describe the histopathological lesions of a new canine
disease characterised by progressive ataxia, head tremor and sei-
zures, and to deduce the cause of the lesions.
METHODS: Formalin-fixed tissues were processed into par-
affin wax and epoxy resin for light and transmission electron
microscopy of variously stained tissue sections.
RESULTS: Significant lesions relevant to the disease were found
only in the brain. They consisted of hypoplasia of the cerebel-
lum and the presence of large pale inclusions in the perikaryon
of neurons in the neocortex and in macrophages. The inclu-
sion material was not compartmentalised and did not stain for
carbohydrate, mucopolysaccharide or lipid. This material dis-
placed nuclei to the periphery of the cells where they were seen
as basophilic distorted crescent-shaped structures.
By 8 months of age, there was also hypermetria of the hindlimbs
associated with reduced conscious proprioception, in addition to
hindlimb ataxia. Sciatic and patellar reflexes were exaggerated.
There was an inconsistent menace response but vision appeared
intact and normal pupillary light reflexes were normal.
Results of haemogram and cerebrospinal fluid (CSF) examina-
tions were normal but the electrolyte panel showed an elevated
chloride concentration of 129 mmol/l (normal range 98–107
mmol/l). A bowel biopsy was negative for lipofuscin-like pig-
ments on histopathological examination.
CONCLUSIONS: The inclusions were probably made of poly-
merised protein similar, though not identical, to those of Pick,
Lewy and Collins bodies that characterise a variety of chronic
neurodegenerative diseases of humans. A genetic basis to this
disease was considered probable.
The owners declined further diagnostic tests such as computerised
tomography and magnetic resonance imaging or brain biopsy,
and elected to continue conservative treatment, as the frequency
of seizures stabilised following increased anticonvulsant dosages.
However, by 15 months of age, seizure activity became more
frequent and did not respond to increased medication. An acute
onset of vomitting occurred associated with abdominal pain and
distension. Exploratory laparotomy revealed a severe necrotising
pancreatitis with secondary peritonitis. Following surgery the dog
was maintained on palliative treatment of pentobarbitone and
intravenous fluids but died shortly afterwards.
KEY WORDS: Dog, ataxia, neurodegeneration, intra-neuronal
inclusions, protein
Introduction
Progressive neurological disease in young dogs with symmetry of
signs and involving several neural systems is highly suggestive of
an inherited metabolic defect, particularly if this includes changes
in learned behaviour or temperament. This report concerns such
a case, which involved a male Cocker Spaniel dog purchased
at 3 months of age. Within a few days of purchase, the new
owners noted he had a clumsy hindlimb gait which was, at first,
attributed to him being a puppy. This became increasingly worse
and, with the onset of head tremor, the dog was presented for
veterinary examination.
The body was placed in a freezer for several hours before be-
ing removed and submitted for necropsy, 15 hours after death;
complete freezing did not occur. Selected tissues, including the
bowel biopsy taken at 6 months of age, were fixed in 10% formol
saline, processed into wax, and sections stained with haematoxy-
lin and eosin (H&E), periodic acid Schiff (PAS), luxol fast blue,
Sudan black, toluidine blue, and alcian blue methods. Formalin-
fixed frozen sections were stained with H&E, PAS and Sudan
black methods. Additional brain neocortex was post-fixed in
1% osmium tetroxide and processed into epoxy resin. “Thick”
sections were stained with toluidine blue for light microscopy,
Case History and Clinical Findings
The dog was subjected to clinical examination on a number of
*
Institute of Veterinary, Animal and Biomedical Sciences, Massey University,
Palmerston North, New Zealand.
CSF
H&E
PAS
Cerebrospinal fluid
Haematoxylin and eosin
Periodic acid Schiff
† Paremata Veterinary Hospital, 15 Paremata Crescent, Porirua, New Zealand.
§ Author for correspondence. Email: R.D.Jolly@massey.ac.nz

204
New Zealand Veterinary Journal 50(5), 2002
Jolly et al
and “thin” sections using the lead citrate/uranyl acetate method
for transmission electron microscopy.
therapy, as vision and pupillary light reflexes appeared normal.
Behavioural changes and seizure activity indicated cerebral
disease. The age of onset and progressive nature of clinical signs
strongly suggested a degenerative central nervous disease, possibly
a lysosomal storage disease. The bowel biopsy was performed to
investigate the possibility of “ceroid-lipofuscinosis with brown
bowel syndrome”, a rare storage disease of Cocker Spaniels but
usually seen in older animals (Jolly et al 1994). Although this case
was negative, a similar case had recently been diagnosed at the
Massey University VeterinaryTeaching Hospital using such means
Pathology
Gross pathology
There was severe necrotising pancreatitis with local fat necrosis,
and a fibrinous peritonitis. The cerebellum was estimated to be
approximately two thirds of normal size.
1
(HM Burbidge , pers. comm.) and provided the rationale for this
Histopathology
diagnostic procedure. The elevated serum chloride concentration
was interpreted to be the result of the potassium bromide this dog
was receiving. Although an ion-specific electrode was used for the
measurement of chloride, this methodology has been reported to
give falsely high chloride values due to the electrode’s inability to
fully discriminate chloride from other halide ions such as bromide
(Trepanier 1995).
With the exception of lesions associated with the pancreas, his-
topathological changes were limited to the brain, in which there
was thrombosis of many blood vessels and some haemorrhage. In
some gyri of the neocortex there were swollen hydropic astrocytes
with pyknotic nuclei, but this was not a generalised lesion. There
were also some superficial freezing artefacts. The architecture of
the cerebellum was relatively normal other than for some slight
segmental thinning of the granular layer. At the tips of three gyri
of the posterior vermis there was a near absence of granular cells
(Figure 1).
The pancreatic necrosis, peritonitis and shock were the immediate
cause of death and were considered unrelated to the underlying
neurological disease. The microscopic vascular lesions were
those of disseminated intravascular coagulation which, together
with the hydropic astrocytes, probably reflected disturbances in
blood clotting and ionic and fluid exchanges associated with the
terminal disease.
Many neurons in the neocortex, particularly in the parietal area,
contained large pale inclusions filling much of the cytoplasm and
displacing normal cytoplasmic structures and the nucleus to the
periphery of the cell (Figure 2). The nucleus was often grossly
distorted, appearing as a thin basophilic crescent-like structure
at one pole of the cell. The pale inclusions appeared opaque,
sometimes slightly basophilic, and frequently showed “chatter”
marks from the microtome knife (Figure 3a). They did not
stain with any of the methods used nor did they fluoresce under
ultraviolet light or rotate polarised light. There were also many
distended cells, interpreted to be macrophages, in which the
nuclei were displaced to the periphery of the cell and, like those in
similarly affected neurons, these were also pyknotic and crescent-
shaped. Although these cells frequently appeared empty, others
contained opaque material with knife “chatter” (Figure 3b), or
stringy remnant material. Some of these cells were seen in close
approximation to neurons in which inclusions were evident, but
they were not seen adjacent to blood vessels, within perivascular
spaces or within the leptomeninges. Subjective assessment of the
density of neurons in the cerebral cortex suggested a loss of
neurons.
The hypoplastic cerebellum noted both grossly and histologi-
cally explains the clinical signs of cerebellar disease. The severe
hypoplastic lesions noted at the tips of three gyri, one of which is
depicted in Figure 1, were not considered artefacts of sectioning.
The large pale intra-cytoplasmic neuronal inclusions probably
reflect the neuronal dysfunction responsible for signs that were
interpreted as being due to cerebral disease. The relationship
between these two distinct changes is uncertain. The opaque-
ness of the inclusion material and knife “chatter” was strongly
reminiscent of that seen in colloid of thyroid follicles, suggesting
a similar consistency. That it was relatively solid is also indicated
by flattening of the nuclei and displacement of organelles. There
was no bilayer membrane evident by transmission electron mi-
croscopy between the accumulated material and cytoplasm con-
taining the main cell organelles, indicating that the accumulated
material was not compartmentalised. Flattening of the nucleus
is an unusual lesion, even in lysosomal storage diseases in which
nuclei may also be displaced to the periphery of the cell, but still
appear rounded.
Electron microscopy
Transmission electron microscopy confirmed displacement
of normal cell structures (Figure 4). The inclusion material
was largely electron-lucent but contained a network of fine,
amorphous, electron-dense material, and many electron-dense
particles interpreted to be ribosomes or degenerated ribosomes
(Figure 3c).
The material within cells that were considered to be macrophages
was similar, though not identical, to that within neurons and was
probably derived from phagocytosis of neuronal debris including
the inclusion material. The fact that they sometimes appeared
empty, or partly empty, suggested that the accumulated material
may have been partly digested and, consequently, dissolved during
the preparation of paraffin sections. Such macrophages would be
expected to occur near blood vessels or within perivascular spaces,
but they were not. If the intracellular material was relatively solid,
as suggested, their migratory ability may have been compromised.
An alternative interpretation is that they were microsatellite cells.
Discussion
The hindlimb ataxia and hypermetria with normal hindlimb
strength suggested cerebellar disease. The inconsistent menace
response most likely reflected the sedative effects of anticonvulsant
The histopathology and ultrastructure of inclusion material in
1
Dr HM Burbidge, Institute of Veterinary, Animal and Biomedical Sciences,
Massey University, Palmerston North, New Zealand.

Jolly et al
New Zealand Veterinary Journal 50(5), 2002
205
a
c
b
Figure 3. (a) Light photomicrograph of the cerebral cortex depicting
a neuron (similar to those in Figure 2) in which the accumulated
material shows knife “chatter” marks (H&E, bar=3.7 μm). (b) A light
photomicrograph of a cell interpreted as a macrophage, showing
similar accumulated material with knife “chatter” marks (H&E, bar=
3.7 μm). (c) Transmission electron micrograph of accumulated material
in a neuron of the neocortex which shows as a loose network of
light-staining amorphous occules which contrast with dark-staining
granular material (72,000x).
Figure 1. Light photomicrograph of the tip of a cerebellar folium
showing near absence of granular cells, but presence of Purkinje cells
(H&E, bar=9.1 μm).
Figure 4. Transmission electron micrograph of
a neuron in the
neocortex showing the material that has accumulated and displaced
the nucleus and cytoplasmic organelles to the periphery of the cell.
This material is not compartmentalised (7,800x).
Figure 2. Light photomicrograph of cerebral cortex showing a group
of neurons with large pale intracytoplasmic inclusions that have
distorted the nuclei and displaced them to the periphery of the cells
(H&E, bar=3.7 μm).

206
New Zealand Veterinary Journal 50(5), 2002
Jolly et al
neurons is highly suggestive of intracytoplasmic accumulation of
poorly soluble polymerised protein. The slight basophilia some-
times noted using light microscopy was likely due to the presence
of ribosomes, as structures resembling ribosomes were evident in
these cells on electron microscopy. There are similarities, but also
differences, to other neuronal cytoplasmic inclusions of specific
accumulated proteins associated with chronic neurodegenerative
diseases of humans. These include Pick bodies in Pick and related
diseases, in which the protein is Tau; Lewy bodies in Parkinson’s
and related diseases, in which the protein is _-synuclein (Goedert
1999); Collins bodies in a familial encephalopathy, in which the
protein is neuroserpin (Davis et al 1999ab) and; hyaline bodies
in familial amytrophic lateral sclerosis, in which the protein is
that of copper/zinc superoxide dismutase (Kakizuka 1998). Pick
disease is just one of many neurodegenerative “tauopathies” (Lee
et al 2001).
milial forms of Pick’s, Parkinson’s and neuroserpin accumulating
diseases tend to be dominantly-inherited traits. If this was so for
the disease in the Cocker Spaniel, then a new mutation would be
probable.
In conclusion, a new type of neurodegenerative disease charac-
terised by progressive ataxia, head tremor and seizures, and novel
intra-neuronal inclusion bodies was diagnosed in a 15-month-old
Cocker Spaniel dog. The inclusion bodies were probably made
of polymerised protein similar, though not identical, to those
of Pick, Lewy and Collins bodies that characterise a variety of
chronic neurodegenerative diseases of humans. A genetic basis to
this disease was considered probable.
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Accepted for publication 28 June 2002