Absolute corneal sensitivity and corneal trigeminal nerve anatomy in normal dogs


Barrett, P.M.; Scagliotti, R.H.; Merideth, R.E.; Jackson, P.A.; Alarcon, F.L.

Progress in Veterinary and Comparative Ophthalmology 1(4): 245-254

1991


The sensory nerves in the cornea of normal dogs was investigated through sensitivity testing with a CochetBonnet aesthesiometer, and histological examination. The mean corneal touch threshold (CTT) — or minimum stimulation to the corneal surface to elicit a blink reflex — for 60 normal dogs was 96 mg/0.0113 mm2 in the central cornea and 116, 148, 154 and 159 mg/0.0113 mm2 in the nasal, temporal, dorsal and ventral corneal regions, respectively. Evaluation by skull type revealed significant differences in the corneal sensitivity values. Dolichocephalic skull types had the most sensitive corneas; the mean CTT was 74, 89, 121, 131 and 135 mg/0.0113 mm2 for the central, nasal, temporal, dorsal and ventral corneal regions, respectively. The mean CTT for mesaticephalic skull types was 93, 113, 151, 153 and 160 mg/0.0113 mm2 for the central, nasal, temporal, dorsal and ventral corneal regions, respectively. Corneal sensitivity was lowest in brachycephalic skulltype dogs; the mean CTT was 141, 150, 170, 178 and 186 mg/0.0113 mm2 for the central, nasal, temporal, dorsal and ventral corneal regions, respectively. A modified gold chloride technique was used to stain the neural elements of 14 canine corneas. Whole flat cornea mounts and sectioned histological mounts revealed a mean of 11.5 major stromal nerve trunks entering the midposterior corneal stroma at various perilimbal sites. Stromal nerve trunks extended to the center of the cornea. Repeated branching of the stromal nerve trunks was identified to the level of a subepithelial plexus, which gave rise to fine nerve bundles that extended into the corneal epithelium. Free nerve endings were noted in the corneal epithelial wing cell layer as terminal expansions. The peripheral cornea was innervated by several smaller nerve fibers that penetrated the limbus at the subepithelial level, at various sites encircling the cornea. These smaller nerves branched rapidly and sent extensions into the corneal epithelial wing cell layer to end in terminal expansions.

ORIGINAL
ARTICLE
Absolute
Corneal
Sensitivity
and
Corneal
Trigeminal
Nerve
Anatomy
in
Normal
Dogs
Paul
M.
Barrett",
Randall
H.
Scagliotti
2
,
Reuben
E.
Merideth
3
,
Paul
A.
Jackson
4
and
Fernando
Lazano
Alarcon'
The
sensory
nerves
in
the
cornea
of
normal
dogs
was
investigated
through
sensitivity
testing
with
a
Cochet-Bonnet
aesthesiometer,
and
histological
examination.
The
mean
corneal
touch
threshold
(CTT)
or
minimum
stimulation
to
the
corneal
surface
to
elicit
a
blink
reflex
for
60
normal
dogs
was
96
mg/0.0113
mm
2
in
the
central
cornea
and
116,
148,
154
and
159
mg/0.0113
mm
2
in
the
nasal,
temporal,
dorsal
and
ventral
corneal
regions,
respectively.
Evaluation
by
skull
type
revealed
significant
differences
in
the
corneal
sensitivity
values.
Dolichocephalic
skull
types
had
the
most
sensitive
corneas;
the
mean
CTT
was
74,
89,
121,
131
and
135
mg/0.0113
mm
2
for
the
central,
nasal,
temporal,
dorsal
and
ventral
corneal
regions,
respectively.
The
mean
CTT
for
mesaticephalic
skull
types
was
93,
113,
151,
153
and
160
mg/0.0113
mm
2
for
the
central,
nasal,
temporal,
dorsal
and
ventral
corneal
regions,
respectively.
Corneal
sensitivity
was
lowest
in
brachycephalic
skull-type
dogs;
the
mean
CTT
was
141,
150,
170,
178
and
186
mg/0.0113
mm
2
for
the
central,
nasal,
temporal,
dorsal
and
ventral
corneal
regions,
respectively.
A
modified
gold
chloride
technique
was
used
to
stain
the
neural
elements
of
14
canine
corneas.
Whole
flat
cornea
mounts
and
sectioned
histological
mounts
revealed
a
mean
of
11.5
major
stromal
nerve
trunks
entering
the
midposterior
corneal
stroma
at
various
perilimbal
sites.
Stromal
nerve
trunks
extended
to
the
center
of
the
cornea.
Repeated
branching
of
the
stromal
nerve
trunks
was
identified
to
the
level
of
a
subepithelial
plexus,
which
gave
rise
to
fine
nerve
bundles
that
extended
into
the
corneal
epithelium.
Free
nerve
endings
were
noted
in
the
corneal
epithelial
wing
cell
layer
as
terminal
expansions.
The
peripheral
cornea
was
innervated
by
several
smaller
nerve
fibers
that
penetrated
the
limbus
at
the
subepithelial
level,
at
various
sites
encircling
the
cornea.
These
smaller
nerves
branched
rapidly
and
sent
extensions
into
the
corneal
epithelial
wing
cell
layer
to
end
in
terminal
expansions.
(Progress
in
Veterinary
Com-
parative
Ophthalmology,
Vol.
1,
No.
4,
1991,
pp.
245-254;
Key
words:
canine,
cornea,
nerve,
corneal
sensitivity,
corneal
nerve,
neuroanatomy,
trigeminal
nerve,
aesthesiometer,
gold
chloride.)
*corresponding
author,
Eye
Clinic
forAnimals,
1717N.
Swan
Road,
Tucson,
AZ
85712;
USA,
2
Sacramento
Animal
Medical
Group,
Carmichael,
CA,
'Veterinary
Referral
Service,
Tucson,
AZ
,'Mesa
Veterinary
Hospital,
Mesa,
AZ
S
University
of
Arizona,
Veterinary
Diagnostic
Laboratory,
Tucson,
AZ
T
he
canine
eye
is
nearly
spherical,
with
an
average
diameter
slightly
greater
than
21
mm.'
The
cornea
consists
of
ap-
proximately
17
percent
of
the
eye-surface
area,
and
approxi-
mately
90
percent
of
the
corneal
surface
can
be
exposed
to
the
external
environment.
2
The
innervation
to
this
large
exposed
245
Table
1.
Test
subject
data
by
skull
type.
Breed
Sex
Age(yr)
STT
OD
STT
OS
Dolichocephalics:
Basset
Hound
M
2
19
14
Dachshund
M
2.25
21
19
Collie
M
1.5
24
17
Afghan
M
3
24
27
German
Shepherd
MC
2
35
32
Labrador
Retriever
MC
1.5
14
17
Sheltie
Collie
F
3
17
17
Bull
Terrier
F
1.5
25
21
Afghan
F
5
19
21
Golden
Retriever
cross
F
0.5
19
17
Doberman
Pinscher
F
0.7
17
14
Akita
F
0.7
17
16
German
Shepherd
F
0.8
18
20
Husky
F
3.7
13
15
Chesapeake
Retriever
F
1.1
30
27
Golden
Retriever
cross
FS
7.6
26
24
Labrador
Retriever
FS
5.5
18
16
German
Shepherd
FS
6.2
17
16
Basset
Hound
FS
3.2
19
19
Dalmatian
FS
4.3
19
17
Mesaticephalics:
Akira
cross
M
1
35 35
Poodle
M
0.9
14
13
Toy
Fox
Terrier
M
9.9
19
18
Yorkshire
Terrier
M
11.5
14
17
M.
Pinscher
cross
M
0.7
16
17
Poodle
M
3.1
19
17
Terrier
cross
MC
1.3
24
23
Toy
Fox
Terrier
F
1.6
17
16
Shepherd
cross
F
1.3
26
21
M.
Schnauzer
F
0.5
23
16
Cocker
Spaniel
cross
F
2.5
17
24
Cocker
Spaniel
F
0.6
18
22
Poodle
cross
FS
10.7
18
17
Shepherd
cross
FS
7.5
14
13
Terrier
cross
FS
10
17
12
Maltese
FS
10
24
16
Scottish
Terrier
FS
6.5
14
11
Beagle
cross
FS
3.5
14
16
Poodle
FS
8.5
19
17
Queensland
Blue
Heeler
FS
3
21
24
Brachycephalics:
Japanese
Chin
M
1
26
29
English
Bull
Terrier
M
4
21
21
Lhasa
Apso
Terrier
M
2.5
21
24
French
Bull
Terrier
M
3
23
24
Japanese
Spaniel
M
2.1
16
24
Lhasa
Apso
M
8
17
24
English
Bull
Terrier
M
0.9
21
25
English
Bull
Terrier
F
2
30
21
Lhasa
Apso
F
0.7
14
16
French
Bull
Terrier
F
0.8
27
25
Japanese
Chin
F
2.7
26
19
Pug
F
2
22
19
Japanese
Spaniel
F
2.5
18
26
English
Bull
Terrier
F
0.3
20
23
Lhasa
Apso
F
0.6
21
19
Lhasa
Apso
F
9
16
17
French
Bull
Terrier
F
2
22
22
Cockapoo
Cross
F
0.7
11
13
Lhasa
Apso
F
2.5
14
16
Boston
Bull
Terrier
F
2.5
20
20
Progress
in
Veterinary
&
Comparative
Ophthalmology
/Volume
1,
Number
4
surface
has
been
previously
unreported
in
the
dog.
Several
reports
of
the
nerve
supply
to
the
cornea
have
been
made
in
other
species,
including
humans,
cats
and
rabbits."
In
these
reports,
it
was
found
that
a
gradation
exists
between
these
species,
with
human
corneas
having
the
greatest
number
of
nerve
fibers
followed
by
lesser
numbers
in
the
cat
and
rabbit,
respectively.
In
addition,
it
was
reported
that
the
cornea
is
innervated
by
branches
of
the
trigeminal
nerve,
which
give
rise
to
a
variable
number
of
large
sensory
nerves
which
penetrate
the
limbus
into
the
corneal
stroma,
progressing
to
the
subepithelial
level
and
terminating
in
the
epithelium
as
nerve-ending
expansions.
Adrenergic
or
sympathetic
nerves
have
also
been
lo-
cated
in
the
cornea
of
various
species
by
specialized
staining
techniques.'
Physical
stimulation
to
a
normal
cornea
initiates
a
reflex
leading
to
a
blink
response,
referred
to
as
the
corneal
blink
reflex.
The
degree
of
stimulation
at
the
corneal
surface
to
cause
a
blink
reflex
has
been
studied
in
several
species
with
a
commercially
available
testing
instrument.
4
'"
A
gradation
also
exists
in
the
sensitivity
of
the
cornea,
which
appears
to
be
inversely
proportional
to
the
number
of
stromal
nerve
fibers
present.
Humans
are
the
most
sen-
sitive,
with
the
cat
and
rabbit
having
less
sensitivity,
respectively.
The
stimulation
for
a
normal
blink
is
stated
to
be
caused
by
changes
in
the
hypertonicity
of
the
tear
film,
which
is
communicated
to
the
epithelial
nerve
endings
through
the
epithelial
tight
junctions.'
Human
corneal
sensitivity
is
altered
in
several
disease
processes
and
is,
therefore,
an
important
piece
of
information
in
the
diag-
nostic
work-up
of
a
patient.
Some
diseases
that
display
decreased
corneal
sensitivity
in
humans
are
herpetic
keratitis,
leprosy,
diabetes,
neurotrophic
keratitis,
and
following
penetrating
keratoplasty.
11
The
degree
of
corneal
stimulation
required
before
the
blink
reflex
occurs
has
not
been
intensively
studied
in
the
dog.
The
purpose
of
this
study
was
to
determine
the
minimal
corneal
touch
stimulation
required
-
or
corneal
touch
threshold
(CTT)
-
to
elicit
the
corneal
blink
reflex
in
the
normal
canine.
Additionally,
the
quantity
and
distribution
of
nerves
in
the
normal
canine
cornea
was
investigated.
A
correlation
was
made
relating
the
corneal
sensitivity
and
corneal
nerve
anatomy
by
comparing
the
determined
CTT
and
regional
corneal
innervation.
Materials
and
methods
Corneal
sensitivity
Sensitivity
testing
was
performed
on
120
normal
eyes
of
60
healthy
normal
dogs,
categorized
according
to
doli-
246
CO
JE
NVERSKA,
TASFUE
dUlt
I
A'
12
100
I
.—
du
t4
wr.
du
rn
dwn
4
V
d
0
men
tir,I,
MrtKN+O
i1
t
Yakut*
rromurd,
an
p
,
0
,,
urau
,
0n
2
,
dv
,
M1040
vaAnps
cd
Wedstored
m
dun
au
no,
‘..
W[00
,
0
0uc0,00010
in
pr
nom)
0
1,
08
6
1,4C,
1.84
2,40
:.2Cc
4.60
f
,!'•
12
12
Progress
in
Veterinary
&
Comparative
Ophthalmology
/Volume
1,
Number
4
Figure
1.
The
Cochet-Bonnet
Aesthesiometer
with
instrument
calibration
chart.
DORSAL
SAL
I
TEMPOR
CENTRAL
VENTRAL
Figure
2.
Schematic
of
the
canine
eye,
demonstrating
the
CTT
test
areas
within
each
corneal
region,
as
indicated
by
small
shaded
ovals
and
small
shaded
circle
centrally.
chocephalic,
mesaticephalic
and
brachycephalic
skull
types,
and
identified
by
age,
breed
and
sex.
Test
subjects
were
considered
normal
and
acceptable
for
normal
corneal
sensitivity
testing
if
they
could
pass
an
ophthalmological
and
a
neuro-ophthalmological
examination.
The
ophthal-
mic
evaluation
included
a
history
review,
which
had
to
be
free
of
previous
ocular
disease;
and
an
ophthalmological
examination
for
evidence
of
current
ocular
disease,
espe-
cially
corneal
abnormalities.
Schirmer
tear
test
(STT)
values
were
measured
and
had
to
be
9
mm
wetting
per
minute
or
greater.'"
The
neuro-ophthalmological
ex-
amination
consisted
of
evaluating
the
integrity
of
cranial
nerves
II,
III,
V,
VI,
VII
and
VIII."
Table
1
summarizes,
by
skull
type,
the
breed,
sex,
age
and
STT
values
for
all
dogs
tested.
The
CTT
was
determined
by
stimulating
corneal
touch
receptors
with
a
Cochet-Bonnet
aesthesiometer,'
and
observing
whether
a
corneal
blink
reflex
was
produced.
The
testing
procedure
was
conducted
in
an
undisturbed,
quiet
environment,
while
the
test
subjects
were
comfort-
ably
standing
or
in
sternal
recumbency,
using
a
minimum
ofhead
restraint
or
manipulation
of
the
periorbital
adnexae.
Both
corneal
surfaces
from
each
animal
were
stimulated
in
prescribed
regions
with
the
0.12-mm-diameter
nylon
monofilament
ofthe
aesthesiometer.
The
nylon
filament
of
the
Cochet-Bonnet
aesthesiometer
can
be
varied
in
length
so
that
the
pressure
applied
to
the
cornea
may
vary
from
11
to
200
mg
per
0.0113
mm
2
(scale
reading
of
6.0
to
0.5
cm)
(Figure
1).
The
cornea
was
divided
into
five
regions:
a
9-
mm-diameter
circular
central
region;
and
four
curvilinear
rectangles,
approximately
11-mm
long
and
5-mm
wide,
encircling
the
limbus
(Figure
2).
The
stimulations
were
delivered
to
the
cornea
with
the
aesthesiometer
in
smaller
defined
areas
within
each
corneal
region.
These
defined
areas
were
approximately
a
5-mm-diameter
circular
area
within
the
central
region,
and
oval
areas
4-mm
long
by
2-mm
wide,
placed
3-mm
central
to
the
limbus
in
the
dorsal,
ventral,
nasal
and
temporal
corneal
regions
(Figure
2).
Testing
began
with
the
aesthesiometer
nylon
monofilament
length
at
4.0
cm,
which
was
advanced
until
the
tip
of
the
extended
monofilament
contacted
the
cornea
and
produced
a
slight
(approximately
4
percent)
filament
deflection
or
bend.
The
length
of
the
monofilament
was
decreased
at
0.5
cm
increments
until
the
animal
demon-
strated
consistent
corneal
blink
reflexes
in
response
to
the
stimulus.
A
minimum
of
five
attempts
to
elicit
a
blink
response
were
made
on
each
corneal
region
at
each
monofilament
length.
When
no
blink
response
was
ob-
served,
shortening
of
the
monofilament
continued
until
a
positive
blink
response
was
noted
to
at
least
half
of
the
stimulations.
When
this
level
of
blink
response
was
dem-
onstrated
by
the
test
subject,
the
corneal
touch
threshold
had
been
reached.
The
length
of
the
nylon
monofilament
when
the
blink
responses
were
elicited
was
recorded
for
each
corneal
region
in
the
following
order:
central,
nasal,
dorsal,
tempo-
ral
and
ventral.
If
the
animal
became
excited,
distracted,
or
otherwise
refractory
during
the
testing
procedure,
the
examination
was
postponed.
Most
CTT
measurements
were
taken
by
one
observer,
thereby
minimizing
a
potential
confounding
variable
(inter-observer
differences
or
varia-
tions
in
testing
technique).
Fluorescein
stain
was
instilled
into
the
conjunctival
sac
of
randomly
selected
dogs,
after
the
CTT
had
been
deter-
mined,
to
reassess
the
integrity
of
the
corneal
epithelial
surface.
The
mean
of
the
recorded
lengths
of
the
247
Table
2.
Mean
regional
corneal
touch
thresholds
for
all
dogs
tested.
Central
Nasal
Temporal
Dorsal
Ventral
Number
of
eyes
Mean
mono-
filament(cm)
Standard
deviation-4-
Mean
CTT
(mg/S)
120
120
120
120
120
1.55
1.31
0.96
0.90
0.85
.54
.49
.34
.35
.29
96
116
148
154
159
S=
0.0113
mm
2
Table
3.
Corneal
touch
thresholds
for
the
dog
by
skull
type
.
Dolicho-
Mesati-
Brachy-
cephalics
cephalics
cephalics
Number
of
eyes
40
40
40
Mean
monofilament
length
in
cm
and
{mg/S}
Central
Nasal
Temporal
Dorsal
Ventral
S=
0.0113
mm'
2.03
{74}
1.58
{93}
1.05
{141}
1.64
{89}
1.35
{113}
.94
{150}
1.23
{121}
.93
{151}
.73
{170}
1.11
{131)
.91
{153}
.67
{178}
1.09
{135}
.86
{160}
.60
{186}
Progress
in
Veterinary
&
Comparative
Ophthalmology
/Volume
1,
Number
4
monofilament
at
threshold
stimulations
were
calculated.
A
plot
of
the
standardized
values
for
the
aesthesiometer
was
used
to
generate
a
curve
representing
the
deflected
filament
tip
pressure
at
any
filament
length.
Extrapolation
from
the
standard
curve
was
used
to
convert
the
CTT
means
to
a
standardized
unit
of
force
(mg/0.0113
mm
2
).
A
detailed
statistical
analysis
was
performed
using
paired
and
un-
paired
Student
t
tests.
Corneal
trigeminal
nerve
anatomy
Fourteen
corneas
were
harvested
from
seven
dogs.
Using
a
stab
incision
at
the
12
o'clock
position,
3
mm
posterior
to
the
limbus,
the
incision
was
extended
along
the
limbal
border
with
fine
iris
scissors,
until
the
cornea
and
attached
limbal
tag
at
the
dorsal
border
(which
served
as
a
reference
marker
for
corneal
position
identification)
was
completely
removed.
The
corneas
were
processed
by
a
modified
gold
chloride
technique'
adapted
for
the
canine
cornea.
Harvested
tissues
were
placed
in
cassettesa
to
flatten
the
corneas,
and
then
bathed
in
10
percent
phosphate
buffered
formalin
acidified
to
pH
5.5
with
HC1
for
five
minutes.
Fixation
was
continued
in
a
1
percent
citric
acid
buffer
solution
at
pH
3.5
for
15
minutes.
The
corneas
were
removed
from
the
cassettes
and
immersed
in
a
1
percent
gold
chloride
solution
for
25
minutes,
and
then
incubated
in
200
ml
distilled
water
acidified
with
12
drops
glacial
acetic
acid
for
12
to
24
hours
at
room
temperature.
Following
incubation,
the
corneas
were
placed
in
70
percent
isopropyl
alcohol
and
immediately
split
into
two
lamellae.
Splitting
of
the
lamellae
was
achieved
by
incising
with
a
64
beaver
blade
under
20X
magnification
through
the
corneal
stroma,
parallel
to
the
endothelial
and
epithe-
lial
surfaces.
The
corneas
were
then
torn
along
the
incised
stromal
lamellae,
resulting
in
an
epithelial/anterior
stromal
section
and
an
endothelial/posterior
stromal
section.
After
separation,
the
lamellae
were
processed
through
graded
alcohol
solutions
and
briefly
placed
in
xylene
for
clearing.
Complete
lamellae
were
mounted
flat
in
permount
be-
tween
two
glass
slides,
and
compressed
for
24
hours
with
elastic
bands.
Four
intact
whole
corneas
were
processed
through
the
same
staining
method,
and
then
fixed
an
ad-
ditional
1
5
minutes
in
acidified
buffered
formalin
before
dehydration
in
the
graded
alcohol
baths
and
xylene.
After
embedding
the
four
corneas
in
tissue
mounting
blocks',
7-µrn
sections
(transverse
and
longitudinal)
were
obtained
and
mounted
using
permount-coated
coverslips.
Results
Corneal
sensitivity
Mean
regional
CTT
values
for
all
dogs
tested
are
given
in
Table
2.
A
significant
and
variable
decrease
in
corneal
sensitivity
was
identified
when
comparing
the
central
vs.
peripheral
corneal
regions
(P<0.0001).
The
central
region
was
the
most
sensitive,
followed
by
the
nasal,
temporal,
dorsal
and
ventral
regions,
respectively.
The
nasal
and
temporal
corneal
regions
had
significantly
higher
corneal
sensitivities
than
either
the
dorsal
or
ventral
corneal
regions
(P<0.0001).
When
the
CTT
was
compared
against
skull
type,
a
significant
difference
was
identified.
Corneal
sensitivity
was
lower
in
the
brachycephalic
dogs,
as
compared
to
the
mesaticephalic
and
dolichocephalic
dogs.
Table
3
lists
the
mean
CTT
of
the
three
skull
types
for
each
corneal
region
tested.
The
dolichocephalic
breeds
have
a
significantly
higher
sensitivity
than
all
dogs
tested
(P<0.0001).
Dolichoce-
phalic
dogs
have
significantly
higher
sensitivity
than
the
brachycephalics
in
the
central,
nasal
and
dorsal
corneal
regions
(P<0.02).
Dolichocephalic
dogs
are
also
significantly
248
4
1
'
0.5
1.0
1.5
2.0
2.5
25
20
15
10
1
m m
Progress
in
Veterinary
&
Comparative
Ophthalmology
/Volume
1,
Number
4
Number
of
eyes
Monofilament
length
in
centimeters
Brachycephalics
Mesaticephalics
0
Dolichocephalics
Figure
3.
Distribution
of
central
corneal
sensitivity
based
on
skull
type.
more
sensitive
than
the
mesaticephalics
in
the
central
and
dorsal
corneal
regions
(P<0.01).
The
brachycephalic
breeds
have
significantly
higher
CTT
in
all
corneal
regions
than
all
dogs
tested
(P<0.04).
Figure
3
illustrates
the
relative
distribution
of
central
corneal
sensitivity
levels
for
each
canine
skull
type
and
demonstrates
the
higher
sensitivity
level
of
the
longer
muzzled
dog.
No
significant
sex
differences
were
detected
in
corneal
sensitivity
in
the
cumulated
data
(P>0.47),
or
by
skull
type
(P>0.07-0.45).
No
significant
difference
was
detected
between
left
and
right
eyes
for
all
dogs
tested
(P>0.69),
or
between
skull
types
(P>0.43-0.71).
Classification
of
the
test
subjects
by
age
was
performed
with
one
group,
6
years
or
less
(n=48),
and
the
second
group,
over
6
years
of
age
(n=12).
No
significant
difference
was
detected
based
on
age
of
those
tested
(P>0.11);
however,
80
percent
of
the
population
of
dogs
tested
were
less
than
6
years
of
age.
Corneal
trigeminal
nerve
anatomy
The
canine
cornea
is
innervated
by
a
mean
of
11.5
1SD)
large
stromal
nerve
trunks
(range
=
10
to
14).
These
nerve
trunks
entered
the
mid-stromal
corneal
region
ar
the
limbus.
The
nerves
entered
at
various
sites
around
the
circumference
of
the
cornea,
and
no
consistent
pattern
or
specific
limbal
region
through
which
the
nerves
consis-
tently
entered
the
corneal
stroma
was
identified.
These
stromal
nerves
passed
radially
into
the
central
corneal
region,
where
they
branched
at
multiple
points,
sending
finer
nerve
fibers
to
the
more
superficial
corneal
regions.
In
Figure
4,
a
large
radially
oriented
nerve
trunk,
with
a
few
J
'
Ott
rrt
.r
Figure
4.
Large
stromal
nerve
originating
from
the
limbus
and
branching
as
it
progresses
to
the
central
region
of
the
cornea
(small
arrows).
Modified
gold
chloride
technique,
X28
magnification.
Figure
5.
Stromal
nerve
trunks
originating
in
close
approximation
from
the
limbus
(large
arrows).
These
were
counted
as
one
stromal
nerve
trunk
that
had
divided
in
the
limbus.
Modified
gold
chloride
technique,
X20.
smaller
branches,
was
considered
a
stromal
nerve.
Occa-
sionally,
two
smaller
stromal
nerves
were
noted
to
originate
from
the
limbal
border
in
very
close
proximity,
as
dem-
onstrated
in
Figure
5.
These
were
counted
as
one
stromal
nerve
trunk
that
had
possibly
branched
in
the
limbal
249
Progress
in
Veterinary
&
Comparative
Ophthalmology
/Volume
1,
Number
4
region.
Some
stromal
nerves
were
noted
to
extend
beyond
the
geographic
center
of
the
cornea.
These
nerves
overlapped
with
other
stromal
nerve
fibers
before
completely
branching,
as
shown
in
Figure
6.
In
some
sections
near
the
center
of
the
cornea,
the
large
stromal
nerves
were
noted
to
combine
with
other
stromal
nerve
fibers
(Figure
7).
In
superficial
planes
of
focus
of
the
central
corneal
regions,
the
small
nerve
trunk
divisions
of
the
deep
stromal
nerves
continued
to
branch
into
fine
fibers.
These
smaller
nerve
bundles
ran
in
the
subepithelial
region
as
a
nerve
plexus.
They
were
noted
to
run
parallel,
with
extensions
into
the
corneal
epithelium
(Figure
8),
and
had
further
branching
and
interconnections
between
nerve
fiber
bundles
(Figure
9).
The
extensions
from
the
subepithelial
plexus
branched
markedly
as
the
nerve
fibers
intertwined
between
corneal
epithelial
cells
(Figure
10).
The
corneal
epithelium
was
richly
supplied
with
nerve
endings
that
extended
into
the
wing
cell
layer.
Nerve
endings
intertwined
between
epithe-
lial
cell
borders
and
ended
in
terminal
expansions,
or
boutons
(Figure
11).
Fine
nerve
fibers
branched
repeatedly
in
the
epithelium,
at
times
in
close
proximity
to
the
terminal
expansion
of
the
nerve
(Figure
12).
The
periphery
of
the
cornea
was
innervated
by
many
smaller
nerves
that
extended
into
a
region
2
to
4
mm
central
to
the
limbus.
These
nerves
were
located
in
more
superficial
planes
of
focus
than
the
deeper
large
stromal
nerve
trunks.
In
the
perilimbal
region,
several
fine
nerve
fibers
entered
the
cornea
from
the
limbus
and
rapidly
split
into
smaller
nerve
fiber
bundles.
The
nerve
bundles
projected
through
the
subepithelial
regions,
branched,
interconnected
re-
peatedly,
and
sent
free
nerve
endings
between
epithelial
cells.
The
methods
of
the
present
study
precluded
a
determi-
nation
of
whether
the
superficial,
perilimbal
nerves
origi-
nated
as
branches
of
deep
stromal
nerves,
or
as
continua-
tions
of
conjunctival
nerves.
Discussion
Corneal
touch
thresholds
and
corneal
sensitivity
are
inversely
proportional;
therefore,
the
higher
the
CTT,
the
lower
the
corneal
sensitivity.
The
mean
central
corneal
touch
threshold
for
the
dog
was
96
mg/0.0113
mm
2
,
but
varied
by
skull
type
(range,
74
mg
for
dolichocephalic-skull
dogs
to
141
mg
for
brachycephalic-skull
dogs).
This
is
a
higher
CTT
than
reported
for
other
species,
including
humans
(10-14
mg),
9
cats
(43
mg),
4
Dutch
pigmented
rabbits
(45
mg),
17
and
albino
rabbits
(87-167).
17
The
ca-
Figure
6.
Stromal
nerve
trunk
branches
in
the
center
of
the
cornea.
Note
the
extensive
nerve
branch
density
and
substantial
overlapping.
Modified
gold
chloride
technique,
X51
magnification.
nine
cornea
exhibited
a
regional
variation
in
sensitivity
similar
to
other
species,
with
the
greatest
sensitivity
level
confined
to
the
central
region.
In
humans,
corneal
sensitivity
is
lower
after
sleeping
because
of
decreased
surface
oxygen
tension
during
ex-
tended lid
closure.
Lower
sensitivity
of
the
canine
ventral
corneal
region
may
be
due,
in
part,
to
the
hypoxic
effect
described
in
people,'
since
the
lower
lid
and
third
eyelid
invariably
contact
the
ventral
corneal
surface.
This
decrease
in
ventral
corneal
sensitivity
is
in
corroboration
with
similar
findings
reported
for
the
cat.
An
earlier
study
examined
the
mechanical
threshold
for
the
cornea
reflex
of
laboratory
animals,
which
included
six
dogs.
18
The
reported
level
for
the
central
cornea
was
4.5
and
4.3
g/mm
2
for
the
right
and
left
eyes,
respectively.
Conversion
of
the
central
corneal
values
from
this
earlier
study
to
mg/0.0113
mm
2
produces
a
value
lower
than
determined
with
the
more
modern
Cochet-Bonnet
aesthesiometer.
This
early
report
described
decreased
pe-
ripheral
corneal
sensitivity,
which
was
substantiated
by
the
present
study.
Canine
regional
CTTs
have
not
been
previously
re-
ported.
The
present
study
demonstrated
significant
differences
in
corneal
sensitivity
between
the
skull
types
and
the
various
regions
of
the
cornea.
Whether
this
repre-
sents
a
difference
in
corneal
nerve
anatomy
or
epithelial
nerve
ending
density
between
the
various
skull
types
remains
to
be
demonstrated.
Very
significant
differences
in
corneal
sensitivity
have
been
documented
in
pigmented
and
albino
rabbits;
however,
the
total
stromal
nerve
num-
ber
and
stromal
nerve
index
were
not
significantly
different
between
these
groups.'
Although
albinos
are
not
exactly
250
e.
25
um
1
30
pm
Progress
in
Veterinary
&
Comparative
Ophthalmology
/Volume
1,
Number
4
IOW
4i7
1.
...
125
pm
0
1
,
Figure
7.
Interconnected
large
stromal
nerve
branches
near
the
central
cornea
(small
arrows).
Modified
gold
chloride
technique,
X100
magnification.
ti
4
240
pm
I
111
1•
*
Figure
10.
Termination
of
a
subepithelial
nerve
plexus
bundle
with
several
branches
extending
into
the
corneal
epithelium.
Note
the
larger
subepithelial
plexus
nerve
bundle
(large
arrow).
Multiple
branches
extend
between
the
corneal
epithelial
cells
(small
arrows)
to
end
in
terminal
expansions.
Modified
gold
chloride
technique,
X500
magnification.
Figure
8.
Branched
corneal
nerves
in
the
subepithelial
plexus.
Small
nerve
Figure
11.
Branching
corneal
epithelial
nerves
(small
arrows)
with
terminal
fibers
ran
in
parallel
orientation
(arrows)
with
extensions
into
the
corneal
expansions
(large
arrows).
Modified
gold
chloride
technique,
X500
epithelium.
Modified
gold
chloride
technique,
X500
magnification.
magnification.
30
Fun
IP
.4
4
..
.•
40
pm
Figure
9.
Interconnecting
subepithelial
plexus
nerve
fibers
(arrows).
Modified
gold
chloride
technique,
X490
magnification.
Figure
12.
Transverse
oblique
section
of
corneal
epithelial
nerve.
Note
nerve
branching
(small
arrows)
very
near
terminal
expansion
of
the
nerve
(large
arrows).
Modified
gold
chloride
technique,
X500
magnification.
251
Progress
in
Veterinary
&
Comparative
Ophthalmology
/Volume
1,
Number
4
like
normally
pigmented
individuals
in
many
ways,
this
rabbit
study
concurred
with
a
study
in
albino
humans.'
9
Because
the
CTT
differences
were
not
attributable
to
cor-
neal
nerve
density
differences,
it
appeared
likely
that
the
CTT
differences
were
attributable
to
some
higher
mecha-
nism
of
the
sensory
system,
subserving
the
sensitivity
of
the
cornea.'
This
higher
mechanism
theory
may
hold
true
for
the
difference
in
corneal
sensitivity
between
blue-eyed
and
brown-eyed
humans,"
and
may
be
applicable
to
the
significant
differences
between
the
skull
types
of
the
pre-
sented
canine
subjects.
No
significant
difference
was
measured
between
the
sexes
of
each
skull
type,
or
between
the
left
and
right
eyes;
this
finding
contradicts
results
reported
in
the
earlier
labo-
ratory
animal
study.'
In
animals,
corneal
sensitivity
testing
with
an
aesthesiometer
provides
a
subjective
evaluation
of
an
ob-
jective
measurement.
Much
debate
has
occurred
in
the
human
literature
regarding
the
classification
of
aesthesi-
ometry
as
an
objective
vs.
subjective
measurement.'
Ob-
jective
methods
monitor
whether
the
blink
reflex
occurs,
while
subjective
measures
are
based
on
cortical
awareness
of
the
sensation
to
the
cornea.
The
tip
of
the
nylon
filament,
when
used
in
humans,
will
cover
four
to
10
epithelial
cells
and
their
nerve
endings,'
leading
to
a
blink
reflex
(objective
measure).
Not
all
blink
reflexes
exhibited
by
human
subjects
reach
cortical
awareness
(subjective
measure).
However,
evaluations
in
humans
found
a
close
correlation
between
objective
and
subjective
measures,
thereby
allowing
the
instrumentation
to
be
used
as
an
objective
tool."
In
the
dog,
the
blink
response
can
occa-
sionally
exhibit
an
incomplete
lid
closure
to
the
corneal
stimulations;
therefore,
periodic
subjective
interpretation
of
the
blink
reflex
must
occur.
Sensitivity
testing
in
animals
is
difficult
for
various
reasons,
including
patient
uncooperation
and
anxiety,
as
well
as
frequent
voluntary
movements
of
the
patient's
globe.
Additionally,
a
margin
of
error
exists
in
the
opera-
tion
of
an
aesthesiometer
when
applied
to
the
dog.
It
has
been
shown
that
the
aesthesiometer
will
deliver
various
pressures
with
changes
in
the
angle
of
bending
of
the
nylon
filament,
at
any
given
filament
length;
and
that
filament
stiffness
is
affected
by
atmospheric
humidity.'
Close
observation
of
filament
flexure,
and
the
observer's
experience
with
the
aesthesiometer
during
its
application
to
animals,
will
de-
crease
and
possibly
eliminate
these
potential
errors.
In
the
dog,
a
blink
reflex
is
the
only
detectable
indica-
tion
that
the
corneal
threshold
has
been
reached.
In
humans,
there
is
variation
in
corneal
sensitivity
under
different
clinical
situations.
Central
corneal
sensitivity
is
slightly
higher
in
room
light
than
under
dark
conditions.
Test
apprehension
in
humans
has
led
to
lid
closure
following
subthreshold
central
corneal
stimulations.'
Loss
of
in-
strument
visualization
and,
thereby,
decreased
patient
apprehension
when
tested
in
darkness
removes
this
margin
of
error.,There
is
also
apparent
cortical
overriding
of
the
blink
reflex
in
humans.
Stimulation
of
the
peripheral
cor-
nea
may
reach
cortical
awareness,
but
does
not
always
elicit
lid
closure."
Whether
test
methodology,
apprehension,
or
cortical
overriding
of
the
stimulations
interferes
with
the
results
in
dogs
remains,
to
be
determined.
Other
nerve
receptors
can
be
stimulatedlp
animals,
also
causing
the
blink
reflex
to
occur.
If
the
operator
inadvertently
touches
the
cilia
of
the
lid
margins,
or
the
pilus
orbitalis
of
the
brow,
a
blink
response
will
occur.
_
Corneal
innervation
of
the
normal
dog
originated
from
two
levels
of
nerves.
A
deep
layer
consisted
of
stromal
nerves
penetrating
the
limbus
at
the
mid-stromal
corneal
level
from
non-specific
points
encircling
the
cornea.
These
nerves
extended
to
the
central
regions
of
the
cornea,
branched
repeatedly
and
progressed
rostrad
to
intertwine
between
epithelial
cells,
innervating
a
majority
of
the
cornea.
A
second
superficial
layer
consisted
of
smaller
nerve
fibers
penetrating
the
limbus
at
the
subepithelial
level
from
various
points
encircling
the
cornea.
These
smaller
nerves
rapidly
separated
into
finer
bundles,
entered
between
the
corneal
epithelial
cells
and
innervated
a
2
to
4
mm
perilimbal
corneal
region.
Corneal
nerve
distribution
in
the
dog
was
very
similar
to
the
other
species
described.
The
dog
averaged
11.5
corneal
stromal
nerves,
which
is
substantially
less
than
reported
for
other
species.
The
human
cornea
contains
30
stromal
nerve
trunks;'
the
rabbit,
12
to
16
stromal
nerve
trunks;
5
and
the
cat,
19
stromal
nerve
trunks.'
The
lower
sensitivity
of
the
canine
cornea
may
be
associated
with
the
decrease
in
nerve
fiber
number.
Whether
a
lower
number
of
stromal
nerve
bundles
results
in
a
reduction
in
superficial
free
nerve
endings
was
not
examined
in
the
present
study.
Nerve
fiber
density
was
greatest
in
the
central
cornea
of
normal
dogs.
The
central
corneal
zone
also
has
the
greatest
exposure
to
the
external
environment;
whereas,
the
perilimbal
regions
are
partially
or
completely
covered
at
any
particular
point
in
time.
The
perilimbal
region
had
a
high
concentration
of
small
nerve
fibers,
however
a
greater
concentration
of
nerve
fiber
numbers
were
present
near
the
center
of
the
cornea.
Increased
central
corneal
exposure
may
explain
the
marked
nerve
density
and
sensitivity
of
that
region.
252
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Conference
Briefs
Optical
Pachymetry
utilizes
the
basic
biomicroscopic
technique
of
specular
reflection.
The
distance
between
the
specular
reflection
of
the
surface
tear
film
and
that
of
the
cornea
is
the
apparent
corneal
thickness.
Several
methods
have
been
used
to
measure
this
apparent
corneal
thickness.
Early
optical
pachymeters
consisted
of
a
micrometer
(Ulbrich's
drum)
on
a
slit-lamp
biomicroscope,
which
measured
the
displacement
of
the
slit
lamp
as
the
focal
point
was
changed
from
the
epithelial
surface
of
the
cornea
to
the
endothelial
surface.
Displacement
of
the
slit
lamp
corresponded
to
corneal
thickness.
Other
early
optical
pachymeters
consisted
of
a
micrometer
scale
in
the
eyepiece
of
the
biomicroscope,
which
was
used
to
measure
the
apparent
thickness
of
the
slit-beam
optic
section
of
the
cornea.
Modern
optical
pachymeters
consist
of
image
doubling
devices
also
attached
to
slit-lamp
biomicroscopes.
(B.C.
Gilger,
S.
A.
McLaughlin,
R.D.
Whitley,
"Corneal
Pachymetry,"
from
the
Program
ofthe
Twenty-First
Annual
Meeting
of
the
American
College
of
Veterinary
Ophthal-
mologists,
October
1990.)
254