J. Anat.

15 Pages·2005·3.3 MB·English

319
J.Anat.(1994)184,pp.319-333,with12figures PrintedinGreatBritain
Dimensions of individual alpha and motor fibres in the
gamma
ventral funiculus of the cat spinal cord
C. FABRICIUS, C.-H. BERTHOLD AND M. RYDMARK
Department ofAnatomy, University ofGoteborg, Sweden
(Accepted23 September 1993)
ABSTRACT
Using light and electron microscopy, axon diameter, myelin sheath thickness (measured as number ofmyelin
lamellae) and internodal length ofalpha and gamma motor axons ofthe L7 ventral root and spinal cord
segment were investigated in serial cross-sections. The CNS internodes ofthe alpha motor fibres had, on
average, an axon diameter of 8.6 gim, 105 myelin lamellae and a length ofabout 560 gim. The CNS
internodes ofthe gamma motor fibres had, on average, an axon diameter of 3.4jm, 66 myelin lamellae and
a length ofabout 440 gim. Axon diameter at the nodes ofRanvier was 30-40% ofthe internodal axon
diameter. Axon diameter, number ofmyelin lamellae and internodal length varied considerably between
consecutive internodes. Statistical analysis showed no systematic increases or decreases. Regression analyses
ofthe scatter plots ofthe number ofmyelin lamellae and internodal length against axon diameter showed
large variations and correlation coefficients ofr < 0.50. In conformity with ventral root (PNS) internodes
(Nilsson & Berthold, 1988) the plotting ofintrafunicular (CNS) internodal myelin volume against intemodal
axon mantle area showed linear correlations with correlation coefficients ofr > 0.90. The mean axon
diameter ofthe investigated CNS internodes was similar to, the mean number ofmyelin lamellae somewhat
lower than, and the mean internodal length considerably shorter than that ofinternodes ofaxons ofthe L7
ventral root (Nilsson & Berthold, 1988). In contrast to the ventral root, the intrafunicular alpha motor fibres
had higher g values (axon diameter/fibre diameter value) and lower illdratios (internodal length/axon
diameter ratio) than is considered optimal for conduction. We consider that these deviations from the
theoretical optimum are not large enough to impair the conduction properties ofthe CNS parts ofthe
motor axons in a significant way.
Key words: Axons; myelin; nodes ofRanvier; internodal length; cat.
Several ultrastructural morphometric studies of
INTRODUCTION
myelinated fibres in the CNS of mammals include
Three structural variables, axon diameter (d), myelin records ofinternodal length and axon diameter(Hess
sheath thickness (nl= number of myelin lamellae) & Young, 1952; Haug, 1967; McDonald & Ohlrich,
and internodal length (id), define the functional 1971; Gledhill et al. 1973; Ohlrich & McDonald,
anatomy ofa myelinated nerve fibre. These variables 1974; Czarkowska etal. 1976; Gledhill & McDonald,
havebeen studied extensively inthePNS ofadult and 1977; Murray & Blakemore, 1980), some include
developing mammals usingultrastructural techniques records ofmyelin sheaththickness and axondiameter
(Fraher, 1978a,b;Mooreetal. 1978;Arbuthnottetal. (Fraher, 1976; 1978a; Berthold & Carlstedt, 1977b;
1980; Friede & Bischhausen, 1980, 1982; Friede et al. Hildebrand & Hahn, 1978; Fraher & Kaar, 1985;
1981; Ritchie, 1982; Berthold et al. 1983; Friede & Biedenbach et al. 1986) and one (Fraher, 1978b)
Beuche, 1985; Fraher & Kaar, 1986; Fraher et al. includes records of all 3 variables. Records ofwhole
1988; Nilsson & Berthold, 1988; for reviews, see and consecutive CNS internodes ofadult animals do
Waxman, 1978, and Behse, 1990). not seem to be available in the current literature.
Correspondence to Dr C. Fabricius, Section ofNeuroanatomy, Department ofAnatomy, University ofGoteborg, Medicinaregatan 3,
S-413 90 Goteborg, Sweden.
320 C. Fabricius, C.-H. BertholdandM. Rydmark
One aim of this study was to measure the 3 Table 1a. Measuredandcalculatedvariablesof2PNSalpha
fundamental fibre variables in consecutive internodes fibre internodes and of 3 CNS alpha fibre internodes
(CNSI-3)
ofthe intrafunicular part ofalpha and gamma motor
fibres. Another aim was to compare the quantitative
PNS' CNS1 CNS2 CNS3 PNS2
morphology of 2 levels of alpha and gamma motor
fibres: theintrafunicularpartoftheCNSlevelandthe Node ofRanvier
ventralrootpartofthePNSlevel(Nilsson&Berthold, Diameter(gim)
n 182 152 48 6
1988).
Mean 3.0 3.3 3.0 2.8
S.D. 0.6 0.7 0.5 0.6
Internode
MATERIAL AND METHODS
Diameter (gm)
Preparativeprocedure n 190 192 176 62 106
Mean 8.8 8.6 8.7 9.2 9.0
Four adult cats (aged 0.5-1 y) and 4 newborn kittens S.D. 1.6 1.6 1.9 2.0 1.4
Myelin lamellae (nl)
wereperfusion-fixed throughtheheartwithaprimary
n 15 189 149 8 106
fixative containing 5% glutaraldehyde in phosphate Mean 129 106 105 88 121
buffer. After the perfusion, the L7 spinal cord S.D. 17 26 24 29 14
segments and the proximal parts of the L7 ventral Internodal length(gm)
n - 141 47 6 106
roots were removed, immersion-fixed for 3 h in the
Mean 593 489 269 1378
primary fixative, rinsed in buffer and cut in 1 mm S.D. 269 161 78 160
thick sagittal slices. The specimens were then post- gvalue
n 189 147 3 106
Mean 0.82 0.82 0.86 0.75
S.D. 0.05 0.05 0.02 0.03
illdratio
n - 141 45 6 106
Mean 73 67 41 158
S.D. 39 22 15 32
Myelincross-section area (jm2)
n 189 146 4 106
Mean 33.3 32.9 28.7 58.0
S.D. 8.8 8.5 14.2 12.7
Axonmantle area (jm2)
n 141 45 6 106
Mean 18731 1:3748 6499 48298
S.D. 8394 5996 1563 9887
Myelin volume (jLm3)
n 141 45 1 106
Mean - 20400 1'5142 4491 80117
S.D. - 11346 7608 0 20094
PNS1 and CNSI-3 include pooled values from all 3 animals of
thepresent study.
PNS' includes records ofthe transitional node. PNS2 includes
pooled datafrominternodesofL7ventral rootsof2cats, Iy and
1 yold (Nilsson & Berthold, 1988).
fixed in osmium tetroxide, rinsed, dehydrated and
embedded in Vestopal W (see Berthold, 1968, and
Carlstedt, 1977, for details regarding the preparatory
Ventral White Grey protocol). The thickness oftheventral funiculus (IFL
rootlet matter matter
inFig. 1)wasmeasuredinallanimalsinapreparation
IFL microscope.
Fig. 1. Schematic drawing ofan L7 spinal cord segment (sagittal SemithinandultrathinsectionwerecutfromtheL7
view). Rings 'a'-'d' refer to photographs in Figure 2. The axons rootlet and its intrafunicular continuation from 3 of
were traced over 1-4 internodes. An axon with 3 internodes was
theadultanimals. Theproximal 500 g.moftheventral
divided into 9parts: (1)PNSinternode; (2)Transitional node; (3)
CNSinternode 1; (4)CNS node 1; (5) CNSinternode2; (6) CNS rootlets were sectioned transversely to the spinalcord
node 2; (7) CNS internode 3; (8) CNS node 3; (9) motoneuron surface. At the surface ofthe spinal cord (see Fig. 1)
soma. IFL,distancebetweentheroot-spinalcordjunctionandthe
the plane ofsectionwas adjusted so as topreserve the
ventral horn (greymatter).
Morphometry ofCNS motorfibres 321
Table 1b. Measured and calculated variables of 2 PNS sectioning, and from the number of sections yielded
gammafibreinternodescndof3 CNSgammafibre internodes by a mesa (for details about the serial sectioning
(CNSI-3) technique, see Berthold et al. 1982b). The semithin
nNS1 CNS2 CNS3 PNS2 sections were placed on glass slides and stained with
PNS1 C
toluidine blue andphotographed at amagnification of
Node ofRanvier x 330. The ultrathin sections were picked up on
Diameter (gim) Formvar coated single-hole copper grids, treated with
n 74 70 31 4
Mean 1.3 1.4 1.3 1.3 uranyl acetate and lead citrate and examined and
S.D. 0.4 0.4 0.4 0.4 photographed in a Philips 400 electron microscope.
Internode The magnification of each photograph was deter-
Diameter (gm)
n 98 89 72 42 91 mined by a standard calibration grid.
Mean 3.4 3.4 3.9 3.9 2.3 In each section series, the axons of a ventral root
S.D. 0.8 0.8 1.0 1.2 0.6 fascicle containing about 100myelinated axons (Table
Myelin lamellae (nl) 1) were identified and numbered. The axons were
n 10 84 72 4 91
Mean 83 70 63 53 53 traced, where possible, from the proximal end of the
21 17 17 21 ventral root through the ventral funiculus into the
S.D. 12
Internodal length (gm) grey matter (Fig. 2). A schematic drawing was made
n 65 36 6 91
Mean 465 408 340 593 ofthe longitudinal view ofeach axon. In this drawing
198 146 165 220 therelative position ofeach semithin section aswell as
S.D.
gvalue the section thickness were known. The distance
n 84 66 0.81 0.65 between the transitional node and each of the CNS
Mean
0.05 0.05 0.05 0.04 nodes along the investigated axons could thus be
S.D.
illdratio determined (Fig. 1).
n 65 36 6 91
Mean 133 117 98 255
53 42 35 67 Measured variables
S.D.
Myelin cross-section area (gm2)
n 84 66 3 91 Axon diameter. The diameter of the axon was
Mean 11.0 10.5 2.6 5.4 estimated at 2 principal levels: along the stereotype
S.D.
Axon mantle area ('m2) part of an internode (d) and at the node of Ranvier
n 65 36 6 91 (d.). The diameter of the axons was calculated from
Mean 7287 6364 5240 5736 measurements of axonal cross-sectional area (in the
S.D. 44037 3193 3262 3257 following, d(a) and dn(a)) and ofaxonal circumference
Myelinvolume(gm3)
n 65 36 2 91 (in the following, d(c) and df(C)) made in light
Mean 5878 4356 5996 5439 micrographs at a final magnification of x 3000 using
S.D. 33887722____33224433 ___773388 __5_5440055 a personal computer equipped with a graphic tablet
PNS' and CNS1-3 inclade pooled values from all 3 animals of and a morphometric program. Paranodal fibre seg-
ments and fibre levels with prominent incisures of
the present study.
pooPlNeSd1daitncalufdreosmrienctoerrdnsod(olefstohfeLt7ravnesnittriaolnalroontosdeo.fP2NcSat2s,i2ncluadneds dSicahmmeitdetr-'La(dn)terrefmearns,wuenrleesasvositdaetde.d o'tIhnetrewrinsoed,altoaxtohne
8
1 y old (Nilsson & Bertho] mean value of 3-4 well separated d(a)measurements
along the same internode. The internodal diameter of
mode oftransversal s,ectioning ofthe axons through- anaxon, asmeasured in the ventral root, was used to
out the intrafunicular continuation oftherootlet. The distinguish between alpha (d ) 5 gm) and gamma
reorientation of the specimens was guided by the (d< 5 jm) motor fibres. 'Nodal axon diameter', (d.),
position of the ventral root relative the longitudinal was measured at the nodal midlevel.
axis ofthe spinal cornd (method ofsectioning, slightly Number ofmyelin lamellae, 'nl'. The lamellae were
modified, according to Fraher, 1978a). A series of counted in the negatives ofthe electron microscopic
about 4000 transveirse semithin (0.5 gm) sections photographs at 1 randomly selected level per inter-
including short nces ofultrathin sections (about nodal segment (cf. Berthold et al. 1983).
seque
100 nm thick) cut aftter every 100th semithin section Internodal length, 'il'. This variable was measured
obtained from e;ach specimen. Section thickness in the drawings ofthe individual axons.
was
calculated from the height ofthe trimmed mesas Sincenland ilcould notbe obtainedfortheventral
was 1
(15-30 gm) that *e successively prepared during root (PNS) part of the fibres, we used previously
wer
322 C. Fabricius, C.-H. Bertholdand M. Rydmark
r
3
ffm
w16
S
I
S
9..
49
Ke0
2(a) -
I
--
1A "-
4'-.,-E.
a
2(d) yt X =
'Y4 sfD~ ~ T3~W2W 'U~~ I~ J-4
Fig. 2. Lightmicrographsof1 oftheanalysedfascicles.Thefasciclewasobservedintheventralroot(a),intheventralfuniculus(b-c)and
intheventralhorn(d).Dotsindicatetheaxonsoftheanalysedfascicle.SCS,spinalcordsurface;GM,greymatter(ventralhorn);S,neuron
soma. Note that 2 ofthe4indicated fibres observed in (a) have disappeared from the section seriesin(d). Bar, 10gim.
Morphometry ofCNS motorfibres 323
presented data from the middle part ofthe adult cat interdependency between different variables was
L7 ventral root (Nilsson & Berthold, 1988) as a tested using regression and correlation analyses.
complement in several of our diagrams and in the Values ofr were calculated and the hypothesis r = 0
Discussion. was tested at the P = 0.05 level for all plots.
Calculated variables
RESULTS
The calculated variables were based on the measured
General morphology
variables of the presently recorded CNS internodes
and of the PNS internodes taken from Nilsson & The transverse contour of the investigated fibre
Berthold (1988). The g values and il/d ratios were bundles was circular in the proximal part of the
calculated since they are considered to be important rootlets. About two thirds ofthe fibres in the rootlet
for the electrophysiological properties of the fibres were classified as alpha motor fibres and one third as
(see, e.g., Brill et al. 1977). gamma motor fibres (Table 1a,b). The CNS-PNS
1. g value: d(a)/(d(a)+2xnlxk); transitional zone protruded from the spinal cord
2. il/dratio: il/d(a). surface and was thus located in the proximal parts of
The calculated variables also included the myelin the rootlets (see Berthold & Carlstedt, 1977a; Fraher
cross section area, used incalculations correlating the & Kaar, 1986; Fraher, 1991). The transverse contour
axonwiththemyelinsheathinatransversesection,and ofthe fibre bundles gradually changed from circular
the internodal myelin sheath volume and the inter- to elliptic in the ventral funiculus (Fig. 2). The mean
nodal axon mantle area, used in the calculations distancebetweentheroot-spinalcordjunctionandthe
correlatingtheaxonwiththemyelinatingcellforwhole ventral horn as measured in a transverse section
internodes. through the L7 segment (IFL in Fig. 1) was 1092 gim
3. Myelin cross-section area: MyCA = 7xnlx forthe adultcats (n = 4) and 217 gm forthenewborn
k(d(C)+kxnl); kittens (n = 4). The 3 examined fascicles measured
4. Internodal myelin sheath volume: MyV= 1189, 1247 and 1328 jm, respectively, from the root-
MyCA x il; spinal cord junction to the grey matter.
5. Internodal axon mantle area: AxMA = d(c) x The number of fibres confined within a fascicle
7i xil; diminished with the distance proximal to the root-
k = 0.009 in the CNS (the CNS myelin period is spinal cord junction (Figs 1, 2). The loss caused by
9.4 nm after preparation; see Hildebrand & Muller, gradual relocation of individual fibres from the
1974); k = 0.012 in the PNS (the PNS myelin period fascicles into the surrounding tracts of the ventral
is 12 nm after preparation, see Finean, 1953). funiculus made it impossible for us to record more
Calculations ofassumed 'fresh state' variables, i.e. than a few ofthe initially identified fibres in the grey
calculations including the compensation for prepara- matter (cf. Fraher et al. 1988). Of the 288 fibres
tory artifacts, were initially performed according to identified in the 3 ventral root fascicles, 245 could be
Berthold et al. (1982a) but were finally deleted since recorded atalevelof400 ,umcentraltotheroot-spinal
the 'fresh state' compensation only had a marginal cordjunction, 181 at 800 gim and 49 at 1200 mm.
effect on the calculations and correlations.
Measured variables
Statistics
Internodalaxon diameter. The mean axon diameter
All variables were stored in a database system. ofthepooled material ofinternodes was 8.8 gm (min:
Mathematical operations, graphic displays and stat- 5.1, max: 12.5) for the PNS alpha motor fibres and
istical computations were performed with the 3.4 jm (min: 1.7, max: 4.9) for the PNS gamma
MINITAB, SAS and Stanford Graphics software. motor fibres. In the CNS internodes 1 and 2 of the
The parametric methods were used at a significance alpha motor fibres the mean diameter was 8.6jim
levelof95%. Thedegreeofvariationbetweenanimals (min: 4.7, max: 12.4) and 8.7 jm (min: 4.2, max:
and different parts of nerve fibres was evaluated by 13.0), respectively. In the CNS internodes 1 and 2 of
ANOVA. the gamma motor fibres the mean diameter was
Similarity between values of the same variable in 3.4jim (min: 1.7, max: 5.5) and 3.9jim (min: 2.2,
consecutive internodes of an individual nerve fibre max: 6.4), respectively (see also Table 1a,b). The
was tested using the paired Student's ttest. The diameter ofindividual axons could both increase and
22 ANA 184
324 C. Fabricius, C.-H. Berthold andM. Rydmark
E
E
Qz. 1.z)
0 2 4 6 8 10 12 14 4 6 8 10 12 14
dCNS1 (9m) dCNS2 (jm)
6 6
5
5 5- 0C
0
4 4-I
3 3-
c z
2 2-
iV
1
1-
0
0 I I I I I
I I
0 2 4 6 8 10 12 14 0 2 4 6 8 14
10 12
dCNS1 (jm) dCNS2 (9m)
200
150
E
s-
Uz) 100 cn
z
50
50
0
0 50 100 150 200 0 250 500 750 1000
n/CNS2 (jm) ilCNS2 (jm)
Figs 3-7 show pooled data from all 3 animals. Triangles andcircles indicate gamma and alphamotor fibreinternodes, respectively.
Fig.3.Axondiameterofconsecutiveinternodes.A / linehasbeenindicatedinthediagrams.(a)TheaxondiametersofthePNSinternodes
(dpNs)plotted againsttheaxondiametersofCNSinternodes (dcNsl). (b)Theaxondiameters ofCNSinternodes (dCNsl)plottedagainst
the axon diameters ofCNS internodes 2 (dCNs2).
Fig. 4. The axon diameters of the transitional nodes (dtn) plotted against the axon diameters of CNS internodes (dcNsl). dt.
0.20+0.33xdCNsl, r=0.92.
Fig. 5. The axon diameters ofCNS nodes (dfCNsl) plotted against the axon diameters of CNS internodes 2 (dcNs2). dfCNsl =0.25+
0.35xdCNS2, r=0.91.
Fig. 6. Numberofmyelinlamellaeinconsecutive internodes. A 1/1 linehasbeenindicatedinthediagram.Thenumberofmyelinlamellae
inCNS internodes (n'cNs1) plotted against the number ofmyelin lamellae in CNS internodes 2 (n'cNs2).
Fig. 7. Internodallengthofconsecutiveinternodes.A / linehasbeenindicatedinthediagram.TheinternodallengthsofCNSintemodes
(i1cNsj) plotted against the internodal lengths ofCNS internodes 2 (ilcNs2).
Morphometry ofCNS motorfibres 325
Table 2. Analysis ofvariance ofthemeasuredandcalculated the following examples further demonstrate the
variablesof2ofthe CNSinternodesshownseparatelyfor the magnitude of the variation. One of the alpha motor
alpha and gamma fibres of the 3 individual experimental
fibres, d= 9.4 gim, hadamyelinsheathof160lamellae
animals
in CNS internode 1 and a sheath of 70 lamellae in
Alpha fibres Gamma fibres CNS internode 2. Another fibre, d= 8.2 gim, had
animal number animal number myelin sheaths with 42 (CNS internode 1) and 158
lamellae (CNS internode 2), respectively.
1 2 3 1 2 3
Internodal length. Mean internodal length was
PNS internode against 558 gim(min: 150,max: 1325)fortheCNSinternodes
CNS internode 1 ofthealphamotorfibres and439 gm(min: 150, max:
Internodal diameter n.s. n.s. n.s. n.s. n.s. n.s. 1075) for the CNS internodes of the gamma motor
Transitional node against
fibres (see also Table 1a,b). The plots of internodal
CNS node I
Nodal constriction + + + n.s. n.s. n.s. lengths of consecutive internodes of the same axons
CNS internode 1 against (Fig. 7) and the plots of internodal lengths against
CNS internode 2
internodal axon diameters (Fig. 9) showed a wide
Internodal diameter + + - + + n.s.
Nodal diameter n.s. - n.s. n.s. n.s. n.s. scatter. Forexample, theCNSinternodes 1,2and3 of
Number ofmyelin lamellae - n.s. + - n.s. n.s. one alpha motor fibre (d= 8 jm), measured 750, 425
Internodal length + n.s. n.s. n.s. n.s. n.s.
and350 jm,respectively,whiletheinternodes 1,2and
gvalue + n.s. - + + n.s.
il/dratio - n.s. + n.s. n.s. n.s. 3 ofanother alpha motor fibre (d= 7 jim), were 200,
263 and 375 jm long, respectively. An increment ofil
Two-tailed difference (Student's ttest): -, difference significant with increasing dwas not observed (see Fig. 9).
(P<0.05), PNS >CNS1 >CNS2; +, difference significant (P<
0.05),CNS1 > PNS,CNS2 >CNSI;n.s.,differencenotsignificant
(P>0.05). Calculated variables
g value. The mean g value was 0.82 (min: 0.67,
decrease when the axons were traced from the PNS max: 0.94)forthe CNSinternodes ofthealphamotor
internodes over the consecutive CNS internodes (Fig. fibres and 0.75 (min: 0.61, max: 0.90) for the CNS
3a,b). Statistical analysis showed no systematic internodes ofthe gamma motor fibres (see also Table
increase or decrease which was common for all 3 1a,b). The g values of consecutive internodes along
animals (Table 2). the same fibre showed wide scatter (Fig. 10). Most
ANOVA for the data showed that the variations in alpha motor fibre internodes (66%) had g values
axon diameter of the CNS internodes could not be exceeding 0.8 and most of these internodes (67%)
related to the individual animals. Consequently, the were also shorter than 600 jm (Fig. 11).
records of axon diameters as well as of other illd ratio. The mean il/d ratio was 70 (min: 17,
parameters were pooled for all 3 animals. max: 204) for the CNS internodes ofthe alpha motor
Nodal axon diameter. All axons showed a con- fibres and 126 (min: 42, max: 318) for the CNS
striction atthenodes ofRanvier. Thediameters ofthe internodes ofthe gamma motor fibres (see also Table
alpha motor axons were somewhat larger at the CNS 1a,b). Most CNS alphamotorfibreinternodes (84%)
nodes than at the transitional nodes (Table 2). The had il/d ratios < 100 and a substantial number of
axon diameter of the transitional nodes was, on the these internodes (32%) had il/dratios < 50 (Fig. 11).
average 36% ofthe internodal diameter and the axon
diameter ofthe CNS nodes was, on the average, 39%
Myelin cross-section area, myelin sheath volume
ofthe internodal diameter (Figs 4,5).
and internodal axon mantle area
Number ofmyelin lamellae. The mean number of
myelin lamellae was 105 (min: 28, max: 170) in the The scatterplot of myelin cross-section area against
CNS internodes of the alpha motor fibres and 66 internodalaxondiameterfortheCNSinternodesofthe
(min: 19, max: 142) in the CNS internodes of the alpha and gamma motor fibres showed a correlation
gamma motor fibres (see also Table la,b). Scatter- coefficient of r = 0.82. The plot made for the CNS
plots of the number ofmyelin lamellae against axon internodes ofthe alphamotorfibres had acorrelation
diameter showed a poor correlation (r < 0.50) with a coefficient of 0.36 and the plot made for the CNS
positive increment ofnlwith increasing dvalues (Fig. internodesofthegammamotorfibreshadacoefficient
8). Figure 6 illustrates the variation in the nlvalues of of0.7, both ofwhich are significantly separated from
consecutive CNS internodes along the same fibre and zero at the0.05 level (Table 3). Regression analyses of
22-2
326 C. Fabricius, C.-H. BertholdandM. Rydmark
180 - 1800 -
8 00
°0 0>
160 - 1600 -
140 - 1400 -
120- 1200 -
100 - - 1000-
80 - - 800-
60- 600 -
40 - 400 -
20 - 200-
0-
I I IT I I I I I I I I I I
0 2 4 6 8 10 12 14 0 2 4 6 8 10 12 14
d(im) d(gm)
1.0 -
0.9-
0.8 -
-
z
(.)n 7
1.0- U./ -
11
0.6 -
0.9- 0 0.5 -
I I I I 1
I
0.5 0.6 0.7 0.8 0.9 1.0
9CNS2
0.8-
VW
0.7- vAh I-rwv- V v
V
v
V
vv V v
0.6- v
VT
V V
0.5
I I I I I I I I I I I
0 50 100 150 200 250 300 350 400 450 500
il/d
InFigs8-11, hollowtrianglesandcirclesindicate theintrafunicular(CNS)gammaandalphamotorfibres, respectively(pooleddata,all 3
animals).Thefilled-intrianglesandcirclesindicatethegammaandalphamotorfibres,respectively,oftheL7ventralroot(PNS)of2animals,
0.5 and 1 y old (Nilsson& Berthold, 1988).
Fig. 8. Numberofmyelinlamellae(nl)ofCNSinternodes 1 andPNSinternodesplottedagainsttheinternodalaxondiameters(d)ofCNS
internodes and PNS internodes.
1
Alphamotoraxons CNS: nl= 141-4.02xd, r=0.25,
PNS: nl=83.8+4.20xd, r=0.43.
Gammamotor axons CNS: nl=29.3+11.5xd, r=0.45,
PNS: nl=-13.3+28.7xd, r=0.85.
Morphometry ofCNS motorfibres 327
Table 3. Linear regression analysis ofscatterplots ofthepooledmeasuredandcalculated variables ofboth alpha andgamma
fibres of2 ofthe CNS internodes andof1 PNS internode
CNS internode 1-2 PNS'
a b r a b r
Alpha and gamma fibre
Nodal axon diameter vs internodal axon diameter:
Transitional node vs CNSinternode 1 0.20 0.33 0.92 n.c.
CNS node 1 vs CNS internode 2 0.25 0.35 0.90 n.c.
Myelinvolume vs axon mantle area -2390 1.21 0.93 -4680 1.76 0.98
Alpha fibre*
Number ofmyelin lamellae vs axondiameter 141 -4.02 0.25 83.8 4.20 0.43
Internodal length vs axondiameter n.c. n.c.
Myelin cross-section area vs axon diameter 16.2 2.00 0.36 -10.6 7.65 0.85
Myelin volume vs axonmantle area -2110 1.20 0.89 -6400 1.79 0.88
Gamma fibre*
Number ofmyelin lamellae vs axon diameter 29.3 11.5 0.45 -13.3 28.7 0.85
Internodal length vs axon diameter 159 85.7 0.34 15.8 249 0.71
Myelin cross-section area vs axon diameter -3.74 4.21 0.70 -10.1 7.69 0.89
Myelin volumevs axonmantle area -401 0.8 0.90 -3400 1.54 0.93
* CNSinternode 1 only.
PNS1, data from the L7 ventral roots of2cats, yYand 1 yold(Nilsson & Berthold, 1988).
Linear regression analysis: a, y-intercept in the linearregression; b, slope ofthe linearregression; n.c., no linearregressioncalculated.
internodal myelin sheath volume plotted against
internodal axon mantle area gave almost identical DISCUSSION
resultsforthe2CNSinternodes,bothwithcorrelation
Methodological considerations
coefficients of about 0.90 (Table 3, Fig. 12a).
Regression analyses ofthese variables for both alpha Inthisstudy,CNSinternodeshavebeenreconstructed
and gamma motor fibres gave: MyV =-2280+ onthebasisofcompleteseriesofconsecutiveultrathin
1.20xAxA, r = 0.93, n = 298. and semithin cross sections. The method is time-
Figure 12b shows the correlation between the consuming and requires careful calibration of the
myelinvolume and the axondiameter and Figure 12d ultramicrotome (Berthold et al. 1982a). However, it
the correlation between the axon diameter and the has the greatadvantage ofgivingcomparatively large
internodal axon mantle area. The simultaneous samples offibres ofall sizes. Itwasnotpossibletouse
influence of the axon diameter and the internodal time saving techniques, such as systematic exclusion
axon mantle area on the internodal myelin sheath ofbatches of 10-20 semithin sections, because ofthe
volume is shown in a 3D graph (Fig. 12c). Multiple uncharacteristic features of CNS paranodal regions
regressionanalyses ofthe3variablesinthisplotgave: and because of the narrow node gaps of the CNS
MyV=-1460+1.3 xAxA-265 xd, r = 298. nodes (Hildebrand, 1971). A node could escape
Fig.9.Internodallengths(il)ofCNSinternodes 1 andPNSinternodesplottedagainsttheinternodalaxondiameters(d)ofCNSinternodes
1 and PNS internodes.
Alphamotor axons CNS: ilagainst d, nonlinear equation r=0.20.
PNS: ilagainstd, nonlinear equation r=0.22.
Gammamotoraxons CNS: il= 159+85.7xd, r = 0.34.
PNS: il= 15.8+249xd, r=0.71.
Fig. 10.gvaluesofconsecutiveinternodes. A 1/1 linehasbeenindicatedinthediagram.ThegvaluesofCNSinternodes 1 (gcNsl)plotted
against thegvalues ofCNSinternodes 2 (gcNs2).
Fig. 11.gvaluesplottedagainstilldratioforallCNSandPNSinternodes. Largehollowcircles,CNSalphamotorfibres(d>7 Am);small
hollowcircles, CNS alpha motorfibres (d= 5.0-7.0gm).
Alpha and gamma motor axons CNS: g=0.85-0.001 xilld, r=0.58.
Alpha and gamma motoraxons PNS: g=0.84-0.001 xilld, r=0.76.
328 C. Fabricius, C.-H. Berthold and M. Rydmark
150
m0x- 100 0
x x
E E
~:-
50- 1-N
AxMA(jm2X 103) d(gm)
150 .150
Xo 100 *100 o
x
x
E
E
> 50-
- 50 .
0O
0
I AxMA(gM2X 103)
15/0 AM(mxO
0
5
10
15
0 25 50 75
AxMA(gm2x 103)
Fig. 12. Theopenspheres showtheintrafunicular(CNS)internodesandthefilledspheresshowL7 ventralroot(PNS)internodes(Nilsson
&Berthold, 1988). ThefullanddashedlinesofthediagramsaretheregressionlinesoftheplotsofdatafromtheCNSandPNSinternodes,
respectively.
(12a) Myelin volume (MyV) plotted against internodal axon mantle area (AxMA).
Alpha motor axons CNS: MyV=-1388+1.17xAxMA, r=0.92.
PNS: MyV=-6400+l.79xAxMA, r=0.88.
Gamma motor axons CNS: MyV=-875+0.89xAxMA, r=0.91.
PNS: MyV=-3400+l.54xAxMA, r=0.93.
(12b) Myelin volume (MyV) plotted against axon diameter (d).
(12c) A 3D graph in perspective showing the myelin volume (MyV) ofall CNS and PNS internodes plotted against the internodal axon
mantle area (AxMA) and the axon diameter (d) ofall CNS and PNS internodes. Alpha and gamma motor axons:
CNS: MyV=-1460+1.3xAxA-265xd,r=0.93,n=298.
PNS: MyV=-6900+1.5xAxA-1450xd, r=0.98, n= 197.
Figures 12a, 12b and 12dshow the reflections ofthe scatterpoints in Figure 12c in 3 conventional 2D scatterplots.
(12d) Axon diameter (d) plotted against internodal axonmantle area (AxMA).