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Sprouting and reflex recovery after spinal nerve lesions in cats
EXPERIMENTAL
NEUROLOGY
Sprouting
73.132-749
(1981)
and Reflex Recovery after Spinal Nerve Lesions in Cats
JOCELYN PRENDERGAST,’ MARION MURRAY, AND MICHAEL E. GOLDBERGER~ Department of Anatomy, The Medical College of Pennsylvania, Philadelphia, Pennsylvania I91 29 Received January 14, 198i The contribution of L7 spinal nerve to the innervation of tibialis anterior muscle varies with the anatomic arrangement of the lumbosacral plexus. When the plexus is “prefixed”. L7 shares the innervation of tibialis anterior with L6 and Sl. “Postfixation” of the plexus results in exclusion of L7 from the innervation of tibialis anterior. L5, L6, and S1 and 52 were cut unilaterally in 22 cats. Postoperatively, in cats with pre$xed plexus the tendon reflex of tibialis anterior was initially weak. Methylene blue staining revealed both normal and degenerating nerves to tibialis anterior. The reflex became stronger during the first week. At that time an increased number of motor end-plates per axon was found. In operated cats with postfixed plexus, the tibialis reflex was abolished. No normal (nondegenerating) nerves or motor end-plates were seen. During the second week the abolished reflex returned. Some axons were now seen in the intramuscular nerve of tibialis anterior. They contained abnormal knot-like structures from which increased numbers of fibers extended. Some of these ended in single, normal-appearing motor end-plates. Reflex recovery can thus be associated with sprouting of peripheral nerves of two types: “homonymous” sprouting of nerves already within a partially denervated muscle occurs rapidly (first week) and may mediate increased strength of a weakened reflex; “heteronymous” sprouting of nerves outside the denervated muscle occurs more slowly (second week) and may mediate the return of a reflex initially abolished by the lesion. ’ To whom reprint requests should be addressed: Department of Anatomy, The Medical College of Pennsylvania, 3300 Henry Ave., Philadelphia, Pa. 19 129. * We would like to acknowledge E. Glazer, K. Golden, B. Goren, and G. Grigonis for their help. This reasearch was supported by grants from the National Institutes of Health (NS15893, NS-16556, and NS- 13768). 732 0014-488618 l/090732- 1StsOZ.OO/O Copyright 8 1981 by Academic Press, Inc. All rights of reproduction in any form reserved.
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INTRODUCTION The acute postoperative period after peripheral nerve injury is characterized by paralysis, and followed by atrophy and loss of contractile strength of the denervated muscle. Subsequently recovery of muscle weight and strength of contraction occur (12, 16, 22, 27). One mechanism available to mediate the recovery is regeneration of cut or damaged axons (6, 10, 13, 19). Because axons regenerate at a rate of about l-3 pm/day (9), the time course of reinnervation and recovery due to regeneration is determined by the distance over which the axon must grow. A second mechanism which can mediate recovery is sprouting of collaterals by undamaged axons, elicited by partial denervation of their terminal fields (8, 15,26). Regeneration and sprouting differ in two obvious ways: (i) the central origins of the reacting axons are different, and (ii) the distances that axons must grow are different. The distance over which collateral growth occurs is short and the time course of reinnervation and therefore recovery is also short. Because both sprouting and regeneration result in reinnervation of denervated muscle, both can be supposed to mediate recovery of reflex or other motor function. Due to the difference in time course of sprouting and regeneration, the recovery time should differ depending on which mechanism is involved. If a lesion is made at some distance from the target, early recovery of a function is likely to be the result of collateral sprouting by surviving axons which were undamaged by the lesion. Due to differences in the central origins of the nerve innervating the denervated muscle during sprouting and during regeneration, one might expect that the characteristics of recovery might also differ under these two conditions. Changes in receptive field size properties after sprouting of sensory nerves (7, 18, 26) and in properites of muscle (8, 12, 15, 16) due to motor nerve sprouting have been described. The contribution of peripheral nerve sprouting to recovery of a centrally integrated function, e.g., reflex activity of a partially denervated muscle, is not well understood. An examination therefore of the time course of reflex recovery after peripheral nerve lesions should further clarify the contribution which peripheral sprouting can make to recovery of useful movements. Overlap between projection fields of intact and axotomized nerves is thought to be one of the conditions for stimulating sprouting by intact axons. In the present experiment we examined sprouting in cat hind limb muscles after a lesion of spinal nerves which contribute to the lumbosacral plexus. Muscle and other peripheral tissue were denervated by cutting L5, L6, Sl, and S2 spinal nerves unilaterally. The L7 spinal nerve was always spared. In the peripheral nervous system, overlap of terminal fields of different spinal nerves occurs due to intermingling of spinal nerves within the
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lumbosacral plexus. The degree to which they are mixed shows considerable individual variablility. This variability is referred to as prefixation or postfixation of the lumbosacral plexus. The peripheral distribution of spinal nerves differs according to whether the lumbosacral plexus is prefixed or postfixed (17, 20, 21). In the prefixed class L5 but not S2 contributes to the plexus and in the postfixed class S2 but not L5 contributes to the plexus (Fig. 1.) The amount of denervation in a muscle produced by a spared L7 nerve lesion will therefore depend on the fixation of the plexus (20, 21). For example, the tibialis anterior muscle is innervated by fibers from the spared L7 in cats with a prefixed plexus but is not innervated by L7 in cats with a postfixed plexus (21). Therefore, in spared nerve preparations, tibialis anterior will be completely denervated in postfixed animals but at least partially innervated in prefixed animals. Thus, it was possible to compare the results of partial denervation of a muscle in some animals with complete denervation of the same muscle in other animals. Differences in reinnervation and in reflex recovery between partially and completely denervated muscle were studied. Furthermore we attempted to determine if differences in reflex recovery from the lesion could be related to sprouting elicited from the spared L7 spinal nerve. MATERIALS
AND
METHODS
Twenty-two adult cats of either sex were used. Surgery was carried out under sterile conditions using Nembutal anesthesia. The spinal nerves on PREFIXED
PLEXUS
POSTFIXED
PLEXUS
L5 L6 L7 Sl s2
FIG. 1. Diagram showing the two classes of fixation of the lumbosacral plexus and the site of transection of the spinal nerves which contribute to the plexus. L5, L6, Sl, and S2 were cut and L7 was spared. In the prefixed class, L5-Sl contribute to the plexus but S2 does not. In the postfixed class, L6-S2 contribute to the plexus whereas L5 does not. In the prefixed condition the tibialis anterior muscle is innervated by L7 and in the postfixed condition it is not.
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RECOVERY
one side of the cord were visualized and L5 and L6 spinal nerves were cut after enlarging the intervertebral foramina; Sl and S2 were cut inside the vertebral canal. All these spinal nerves were transected distal to the dorsal root ganglia. The L7 spinal nerve was always spared. Five tendon reflexes in the cat hind limb were examined on the control and experimental sides daily for 4 postoperative weeks (Table 1). The reflexes in the experimental limbs were compared with those on the control side and graded as absent (-), weak ( +- ), strong (+), and/or clonic (c). Reflexes in the experimental limb which were indistinquishable from those on the control side were classified as normal. Reflexes and locomotion were also filmed. Five of the animals were anesthetized (two at 3 days, and one each at 7, 11, and 13 days postoperative) and either tibialis anterior or soleus and the toe flexor muscles on both legs were exposed and injected with 0.15% methylene blue in physiologic saline such that the entire muscle was stained dark blue (approximately 50 ml). Ten minutes after the injection the muscles were removed, cut in strips, placed on moist gauze, and oxygenated 1 h, at 1 liter/min. Each strip was exposed to the oxygen for 0.5 h per side. The muscle was then fixed 24 h in 8% ammonium molybdate at 4°C washed, cut into thin pieces, and squashed between two slides (4). The entire muscle was dehydrated and coverslipped and examined using a light microscope. The slides were projected and the stained nerves, arborizations, and motor end-plates were drawn with the aid of a drawing tube or were photographed. The animals were perfused through the heart with a 0.5% glutaraldehyde and 4.0% paraformaldehyde mixture. Survival periods are TABLE
1
Tendon Reflexes in Cats’ Hind Limbs Tendon reflex
Tap
Muscle(s)
Response
Hamstrings jerk Lateral (LHj) Medial (MHj)
Tendon of hamstring muscles
Biceps femoris and semitendenosus
Slight flexion of lower leg
Knee jerk (Kj)
Patellar tendon
Rectus femoris
Extension of lower leg
Ankle jerk (Aj)
Achilles tendon
Gastrocnemious and soleus
Extension of foot
Tibialis anterior jerk (TAj)
Tibialis anterior tendon
Tibialis anterior
Flexion of foot
Toe jerk (Tj)
Several tendons on plantar surface of toes
Several toe flexors
Flexion of toes
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PRENDERGAST,
shown in Table plexus, and the exposed, drawn, the lesions and
MURRAY,
AND
GOLDBERGER
2. The spinal cord, dorsal and ventral roots, lumbosacral peripheral nerves to the muscles in both hind limbs were and photographed in situ and then dissected out to confirm to determine the fixation of the lumbosacral plexus. RESULTS Behavior
Normal. Table 1 shows the occurrence of tendon reflexes in normal cat hind limbs. Tap of the tendon results in a brief, single response of leg, foot, or toes followed by return to the resting position. Normally the tendon reflexes are bilaterally symmetrical in strength. Spared Nerve. Acute period (I to 2 days postoperative). On recovery from anesthesia, the tendon reflexes in the hind limb contralateral to the lesion appeared to be normal In the partially denervated hind limb of all cats some reflexes were lost and other reflexes were weak compared with reflexes on the contralateral side. The animals were assigned to two groups on the basis of the status of their tendon reflexes. These animals could be assigned to the same two groups based on the fixation of their lumbosacral plexus, as determined by dissection after killing (Table 3). (i) Prefixed class (N = 9): The tibialis anterior and toe jerks were always present, the ankle, lateral hamstrings and knee jerk were always abolished, and the medial hamstrings reflex was present in 2 cats but absent in 7 cats. (ii) Postfixed class (N = 13): The ankle jerk was present, and the tibialis anterior and knee jerks were lost. The toe jerk was present in 8, the medial hamstrings jerk in 5, and the lateral hamstrings jerk in 11 of the cats. Early recovery period (3 to 7 days). (i) Pre$xed class (N = 9): The responses which were present in the immediate postoperative period became progressively stronger. In addition, tapping the region of the patella, presumably including stimulation of the periostium, produced internal or external rotation of the leg and protraction of the hip in 8 of these cats. TABLE
2
Times of Killing after Lesion and Numbers of Cats in Each Class of Fixation of the Lumbosacral Plexus Acute (1 to 2 days)
Early (3 to 7 days)
Chronic (8 to 19 days)
Prefixed
0
1
8
Postfixed
1
4
8
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AND REFLEX
TABLE
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3
The Pattern of Reflexes Which Were Elicited during the Recovery Period in one Cat Whose Lumbosacral Plexus Was Prefixed and Another Whose Plexus Was Postfixed Prefixed”
Postfixed
Acute (1 to2 days)
Early (3 to I days)
Chronic (8 to 17 days)
Acute (1 to2 days)
Early (3 to 7 days)
Chronic (8 to 19 days)
LHj MHj
(-) (-)
(-) C-J
(+) (+I
(-) (-)
(-) (-)
(-) (+)
Kj
(-)
(-)
(+)
C-J
(-)
C--J
TAj
(+-)
(+I
(+)
(-)
C-1
(+)
Aj
C-1
(-)
(*)
(k)
t+. c)
(+I
Tj
(*I
(+I
(+I
(*I
(+)
(+I
Tendon reflex’
” (-), and Hj cat the h For
Absent; (+), strong; (k), weak; (c), clonic. Notice in the prefixed cat that the Aj return, the TAj and Tj become stronger, and the Kj never recovers. In the postfixed TAj and LHj recover, Aj and Tj become stronger, and the Kj never returns. abbreviations see Table I.
These responses, which are not normally seen, are presumed not to be tendon reflexes because tap of the patella tendon alone produced no response. (ii) Postjixed class (N = 12): The ankle jerk, which was weak initially became stronger and then clonic in 3 cats. In 10 of the 12 cats which survived into this period, tap of the patellar region produced the same rotation and protraction responses as in the prefixed class. Chronic period (8 to 19 days). (i) Prefixed class (N = 8): The ankle jerk returned in 3 cats in which it had been absent. The lateral hamstrings jerk returned in 2 cats in which it had been absent. In 3 cats the tibialis anterior jerk, which had been weak, became strong and then clonic. (ii) Postjixed class (N = 8): Of the 8 cats in this group, the lost tibialis anterior returned in 3 and the lost toe jerks returned in three. Summary of Tendon Reflexes
On the day after surgery the pattern of reflex loss was correlated with the fixation of the lumbosacral plexus, In the prefixed class the ankle, knee, and lateral hamstrings jerks were lost, and the remaining reflexes were weakened. By 7 days all weakened reflexes had become stronger. During the following 12 days the abolished ankle and lateral hamstrings jerk re-
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turned. In the postfixed class in the acute period the tibialis anterior and knee jerks were absent in all cats. In the chronic period the abolished toe and tibialis jerks returned in some cats. The pattern of reflexes which were spared were therefore different for the two classes but in both classes (i) the weakened reflexes became stronger; some even became clonic, and (ii) some of the lost reflexes returned within 3 weeks after the lesion. Anatomy
The following description of morphological changes treats only the tibialis anterior in detail. The changes occurring in tibialis were, however, representative of those occurring in soleus and the toe flexors. Normal (unoperated side, N = 5). The intramuscular nerves coursed across and in between the muscle fibers. The large intramuscular nerves contained many axons, were relatively straight, and branched often. The branches always contained at least five axons. The axons also had a characteristic branching pattern and finally splayed out in a terminal arborization. Usually a single, but occasionally two motor end-plates were seen to arise from each preterminal axon. An example of the characteristics of the normal intramuscular nerves, the axons within them, and the motor end-plates is shown in Fig. 2. Spared nerve (N = 5). Early postoperative periods [(3 and 7 days) (N = 3)]: The morphology*of the tibialis anterior intramuscular nerve on the lesion side differed from normal and also differed between pre- or postfixed animals. In both classes, degenerating axons were present and these were swollen and beaded as they coursed across the muscle (Figs. 3, 4). The motor end-plates on the degenerating terminal axons were swollen and clumped (Fig. 4B, C). Prefixed class (N = 1): The intramuscular nerve was characterized by the presence of some normal appearing axons coursing together with the degenerating axons (Fig. 3). Postfixed class (N = 2): All axons and motor end-plates (Fig. 4) in the tibialis anterior muscle were degenerating. Only a few degenerating motor end-plates were still detectable by 7 days after surgery. Chronic period [ ( 11 and 13 days) (N = 2) 1: In both classes there was some degenerating debris remaining in the muscle. The characteristics of the intramuscular nerve again differed betweeen preand postfixed animals. Prefixed class (N = 1): The pattern of branching of the axons in the intramuscular nerve was identical to that of the control side. Frequently, however, as many as six or seven end-plates branched from a single axon, a condition never seen normally. (Figs. 5, 6). PostJxed class (N = 1): There were a few nondegenerating axons in each intramuscular nerve bundle. A division of the intramuscular bundle often resulted in a single fiber coursing across the section of muscle without branch-
FIG. 2. A-photomicrograph showing an intramuscular nerve and one of its branches stained with methylene blue in a normal cat tibialis anterior muscle preparation, The arrow indicates the characteristic branching pattern of the nerve fibers within the intramuscular nerve. X250. B-photomicrograph showing several motor end-plates (stars) arising from the fibers of a normal terminal arborization. Some motor end-plates are out of the plane of focus (double stars) due to the thickness of the squash. X250. Methylene blue stain.
2 \”
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PRENDERGAST,
MURRAY,
FIG. 3. Photomicrograph of tibialis anterior class, during the acute postoperative period (3 degenerative nerve fibers (arrows) as well as within the intramuscular nerve of the muscle.
AND
GOLDBERGER
muscle and nerve from a cat of the prefixed days postoperative). Note the swollen, beaded normal nerve fibers (stars) coursing together X325. Methylene blue stain.
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‘6’ A
FIG. 4. Photomicrographs and projection drawing of tibialis anterior muscle and nerve in a postfixed cat. The tibialis anterior muscle contains only swollen, beaded degenerating nerve fibers in the intramuscular nerve and swollen degenerating motor end-plates. 7 days postoperative. A-a large intramuscular nerve, in which all nerve fibers are degenerating. X150. Methylene blue stain. B-three degenerating nerve fibers, one of which terminates in a degenerating motor end-plate (DMEP). X500. Methylene blue stain. C.-Projection drawing of the degenerating nerve fibers and motor end-plate (DMEP) shown in B through several planes of focus.
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AND GOLDBERGER
FIG. 5. Photomicrograph shows four terminal nerve fibers (a-d), from one (c) of which three motor end-plates emerge. X500. The star indicates the motor end-plate most nearly in focus. Prefixed cat, 1 I days postoperative, methylene blue stain.
ing. Along the course of the axons, knot-like regions were encountered (Fig. 7). These were never seen in normal muscle or in partially denervated tibialis anterior muscle from cats in the prefixed class. It was not possible to distinguish whether these knot-like thickenings were a point of repeated branching or simply a tangle of nerves. Commonly, however, there were more axons on one side of the thickening. Several terminal axons extended
\
-h
MEP
FIG. 6. A terminal nerve fiber: Three-dimensional, photographic reconstruction through several planes of focus. The negatives of photomicrographs taken at different focal planes were stacked upon one another and photographed to produce the positive. This procedure preserved in detail the terminal nerve fiber and the motor end-plates (MEP) which emerge from it. The reconstruction shows a single nerve fiber from which six motor end-plates emerge. X1 12.5. Prefixed cat, 11 days postoperative.
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from the thickening (Fig. 8) each of which ended in a single, normalappearing motor end-plate. Summary
of Anatomy
Morphological changes in the nerve to tibialis anterior muscle varied as a function of postoperative time and the class of fixation to which the lumbosacral plexus belonged. In the prefixed class in the early postoperative stages the nerve contained bundles of normal and degenerating axons. Each normal axon ended in a single motor end-plate. By 13 days there were several motor end-plates from each preterminal axon but the branching pattern of the axons in the intramuscular nerve appeared normal. In the postfixed class, in the early stages only degenerating axons and motor endplates were seen. In the chronic postoperative periods, however, a few axons were seen in the nerve and these contained knot-like thickenings which appeared to be branch points because the number of axons on one side of the thickening always exceeded the number on the other side. Some of these axons ended in a single, normal appearing motor-end plate. General Summary
(Table 4)
In the prefixed class at 3 days after surgery the reflex was weak. The muscle was presumed to be partially denervated because normal and degenerating axons were seen in the same bundle. One, and occasionally two motor end-plates extended from preterminal axons. Multiple motor endplates were seen to arise from single axons. In the postfixed class acutely, the tibialis anterior jerk was absent; no axons were seen in the muscle except those that were degenerating. In the chronic period, in those animals in which the reflex returned some axons were seen in the muscle. These axons contained knot-like structures from which additional axons extended. Some of these axons terminated in single, apparently normal, motor endplates. DISCUSSION Our results show that after transecting L5, L6 and Sl, S2 (sparing L7) spinal nerves, (i) two different distributions of tendon reflexes are seen in the hind limb ipsilateral to the lesion, (ii) some of the reflexes which were weak acutely became stronger and even clonic later, and (iii) some of the reflexes which were abolished for as long as 2 weeks returned. The two reflex patterns seen acutely in the hind limb depended on the class of fixation of the lumbosacral plexus. In both classes, the reflexes which re-
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AND GOLDBERGER
i’-j-L Experimental
C FIG. 7. Branching of an intramuscular nerve. A.--Camera lucida drawing of a control. B.Three examples of camera lucida drawings of branching of the intramuscular nerves containing knot-like regions described under Results. Postfixed cat, 13 days postoperative. C and D-C is a drawing of branching of a nerve containing two knot-like regions and D is the photomicrograph (X350) from which the drawing (C) was made. Methylene blue stain. The arrow is pointing to a distal branch and shows the orientation. 1 and 2 refer to two knot-like regions and P indicates the presumptive parent axon. Postfixed cat, 13 days postoperative.
mained after the lesion became stronger beginning on the third postoperative day. When the muscle in which weak reflexes could be elicited in the acute period was examined histologically, some normal axons were
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745
always present in the intramuscular nerve. The terminal arborization of the intramuscular nerve was characterized by one, and very occasionally two, motor end-plates per axon. When examined in the chronic period, an increased number of motor end-plates was seen emerging from a single axon. The branching pattern of the intramuscular nerves and the axons within it were normal. The increased number of motor end-plates per axon indicated that sprouting had occurred and had reinnervated the partially denervated muscle. The sprouting apparently occurred early in the first postoperative week concomitant with an increase in strength of the reflex. The axonal sprouts presumably arose from axons which normally innervated the partially denervated muscle. We therefore call this “homonymous” sprouting. It is comparable to sprouting which has classically been called collateral sprouting (8) and has been correlated with several aspects of recovery of function after peripheral nervous system lesions, (2, 7, 12, 14, 16, 18, 25-27). In the tibialis anterior in postfixed animals, no tendon reflex could be elicited in the acute stage. When this muscle was examined in the early, areflexic postoperative stage, only degenerating axons and degenerating motor end-plates were encountered. This is consistent with the observation that in cats with postfixed lumbosacral plexi, electrical stimulation of L7 does not elicit contraction of tibialis anterior (2 1). These observations therefore indicate that in the early stages after surgery the tibialis anterior had no innervation from the spared L7 in postfixed animals. In some animals, the lost reflex returned in the chronic periods. When the tibialis anterior was examined histologically after this reflex recovery, a small number of axons were found. These axons were abnormal in containing knot-like regions from which branches appeared to emanate. Some of these branches coursed singly across the tissue without further branching and others penetrated various distances through the muscle singly or in small groups and terminated with single, normal-appearing motor end-plates. Similar abnormal knot-like regions were described in the course of an examination of muscle in patients whose only other distinction was that of mental illness (5). We interpret our observations to mean that axons had sprouted into the tibialis anterior to reinnervate this denervated muscle. Because in such postfixed animals, all nerves to tibialis anterior had degenerated during the areflexic period, the sprouted axons presumably arose from a nerve innervating another muscle. We propose to call this “heteronymous” sprouting. This differs from the classical description of collateral sprouting in that it apparently does not depend on overlap, i.e., the transected and sprouting nerves do not innervate the same muscle preoperatively. Innervation by sprouting or regeneration by a foreign nerve into a zone previously not
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PRENDERGAST,
MURRAY, TABLE
AND GOLDBERGER 4
Time Course and Character of the Changes in the Reflex and in the Nerves of the Tibialis Anterior Muscle after Transection of Ls, L,,, S,, and S2 (L, Spared) Early (3 to 7 days)
Chronic (8 to 19 days)
Reflex
Anatomy of nerves
Reflex
Anatomy of nerves
Prefixed
Weak
Degenerating and normal
Strong
(i) Several motor end-plates per terminal fiber. (ii) Normal branching of intramuscular nerve and nerve fibers.
Postfixed
Absent
All degenerating
Weak
(i) Few fibers in intramuscular nerve. (ii) Gnarled regions along nerve fibers. (iii) Fibers from gnarled zone course futher or terminate with single motor endplate.
supplied by that nerve has been described in the peripheral nervous system (1, 3, 10, 13, 18, 22-24, 28). Thus two different patterns of reflex recovery can be correlated with two different forms of sprouting. Some of the reflexes which were abolished by surgery never returned. One of these was the knee jerk. Tap of the region of the patella, e.g., skin and presumably periostium, however, resulted in abnormal motor responses at the hip beginning at 3 days after surgery. These responses could have resulted from the unmasking of latent or inhibited reflex pathways when the knee jerk was abolished and this process might require 2 or 3 days after the surgery before the reflexes could be elicited. On the other hand, the time course of their appearance suggested that they also might result from axonal sprouting. The present experiment demonstrates that recovery of a reflex, i.e., a centrally integrated function, which occurs in less than 1 month after cutting spinal nerve in cat, must be due, at least in part, to sprouting of axons which survive the lesion. Early “homonymous” sprouting or later “heteronymous” sprouting always resulted in adaptive recovery. For example ( 1l), when the tibialis anterior jerk was absent, the foot always slipped backward during stepping, but when this reflex returned no slipping during stepping was seen.
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B FIG. 8. Terminal nerve fibers as they branch from a gnarled region of an intramuscular nerve. In the photomicrograph (A) and the reconstruction (B) the fibers labeled (a, b, c) and motor end-plates labeled (1, 2, 3) are the same. A-photomicrograph showing two nerves. From one (arrow), three branches arise (a, b, c) and terminate in motor end-plates (1, 2, 3). Notice that parts of the terminal nerves and the motor end-plates are out of the plane of focus due to the thickness of the squash. Postfixed cat, 13 days postoperative, methylene blue stain. B.-this three-dimensional photographic reconstruction was made through several planes of focus by the same method as Fig. 6 and shows the three branches (a, b, c) and the motor endplates (1, 2, 3) which arise from them. The negative for photograph (A) above was 1 of 10 used to make this reconstruction. Notice that the terminal fibers and the motor end-plates (MEP) can be seen in more detail in the reconstruction than in the photomicrograph (A).
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21. SHERRINGTON, C. S. 1892. Notes on the arrangement of some motor fibres in the lumbosacral plexus. J. Physiol. (London) 13: 621-112. 22. SPERRY, R. W. 1945. The problem of central nervous reorganization after nerve regeneration and muscle transposition. Q. Rev. Biof. 20: 31 l-369. 23. STIRLING, R. V. 1973. The effect of increasing the innervation field sizes of nerves on their reflex response time in salamanders. J. Physiol. (London) 229: 657-679. 24. UNGAR-SARGON, J., AND M. E. GOLDBERGER. 1980. Degree of specificity maintained by sprouting and regenerating motor nerves to cat tibialis anterior muscle. Sot. Neurosci. Abstr. 6: 185. 25. VAN HARREVELD, A. 1945. Reinnervation of denervated muscle fibers by adjacent functioning motor units. Am. J. Physiol. 144: 477-493. 26. WEDDELL, G., L. GUTTMANN, AND E. GUTMANN. 1941. The local extension of nerve tibres into denervated areas of skin. J. Neural. Neurosurg. Psychiatr. 4: 206-225. 27. WEISS, P., AND M. V. EDDS, JR. 1946. Spontaneous recovery of muscle following partial denervation. Am. J. Physiol. 145: 587-607. 28. WEISS, P., AND H. HOAG. 1946. Competitive reinnervation of rat muscles by their own and foreign nerves. J. Neurophysiol. 9: 413-418.
NEUROLOGY
Sprouting
73.132-749
(1981)
and Reflex Recovery after Spinal Nerve Lesions in Cats
JOCELYN PRENDERGAST,’ MARION MURRAY, AND MICHAEL E. GOLDBERGER~ Department of Anatomy, The Medical College of Pennsylvania, Philadelphia, Pennsylvania I91 29 Received January 14, 198i The contribution of L7 spinal nerve to the innervation of tibialis anterior muscle varies with the anatomic arrangement of the lumbosacral plexus. When the plexus is “prefixed”. L7 shares the innervation of tibialis anterior with L6 and Sl. “Postfixation” of the plexus results in exclusion of L7 from the innervation of tibialis anterior. L5, L6, and S1 and 52 were cut unilaterally in 22 cats. Postoperatively, in cats with pre$xed plexus the tendon reflex of tibialis anterior was initially weak. Methylene blue staining revealed both normal and degenerating nerves to tibialis anterior. The reflex became stronger during the first week. At that time an increased number of motor end-plates per axon was found. In operated cats with postfixed plexus, the tibialis reflex was abolished. No normal (nondegenerating) nerves or motor end-plates were seen. During the second week the abolished reflex returned. Some axons were now seen in the intramuscular nerve of tibialis anterior. They contained abnormal knot-like structures from which increased numbers of fibers extended. Some of these ended in single, normal-appearing motor end-plates. Reflex recovery can thus be associated with sprouting of peripheral nerves of two types: “homonymous” sprouting of nerves already within a partially denervated muscle occurs rapidly (first week) and may mediate increased strength of a weakened reflex; “heteronymous” sprouting of nerves outside the denervated muscle occurs more slowly (second week) and may mediate the return of a reflex initially abolished by the lesion. ’ To whom reprint requests should be addressed: Department of Anatomy, The Medical College of Pennsylvania, 3300 Henry Ave., Philadelphia, Pa. 19 129. * We would like to acknowledge E. Glazer, K. Golden, B. Goren, and G. Grigonis for their help. This reasearch was supported by grants from the National Institutes of Health (NS15893, NS-16556, and NS- 13768). 732 0014-488618 l/090732- 1StsOZ.OO/O Copyright 8 1981 by Academic Press, Inc. All rights of reproduction in any form reserved.
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INTRODUCTION The acute postoperative period after peripheral nerve injury is characterized by paralysis, and followed by atrophy and loss of contractile strength of the denervated muscle. Subsequently recovery of muscle weight and strength of contraction occur (12, 16, 22, 27). One mechanism available to mediate the recovery is regeneration of cut or damaged axons (6, 10, 13, 19). Because axons regenerate at a rate of about l-3 pm/day (9), the time course of reinnervation and recovery due to regeneration is determined by the distance over which the axon must grow. A second mechanism which can mediate recovery is sprouting of collaterals by undamaged axons, elicited by partial denervation of their terminal fields (8, 15,26). Regeneration and sprouting differ in two obvious ways: (i) the central origins of the reacting axons are different, and (ii) the distances that axons must grow are different. The distance over which collateral growth occurs is short and the time course of reinnervation and therefore recovery is also short. Because both sprouting and regeneration result in reinnervation of denervated muscle, both can be supposed to mediate recovery of reflex or other motor function. Due to the difference in time course of sprouting and regeneration, the recovery time should differ depending on which mechanism is involved. If a lesion is made at some distance from the target, early recovery of a function is likely to be the result of collateral sprouting by surviving axons which were undamaged by the lesion. Due to differences in the central origins of the nerve innervating the denervated muscle during sprouting and during regeneration, one might expect that the characteristics of recovery might also differ under these two conditions. Changes in receptive field size properties after sprouting of sensory nerves (7, 18, 26) and in properites of muscle (8, 12, 15, 16) due to motor nerve sprouting have been described. The contribution of peripheral nerve sprouting to recovery of a centrally integrated function, e.g., reflex activity of a partially denervated muscle, is not well understood. An examination therefore of the time course of reflex recovery after peripheral nerve lesions should further clarify the contribution which peripheral sprouting can make to recovery of useful movements. Overlap between projection fields of intact and axotomized nerves is thought to be one of the conditions for stimulating sprouting by intact axons. In the present experiment we examined sprouting in cat hind limb muscles after a lesion of spinal nerves which contribute to the lumbosacral plexus. Muscle and other peripheral tissue were denervated by cutting L5, L6, Sl, and S2 spinal nerves unilaterally. The L7 spinal nerve was always spared. In the peripheral nervous system, overlap of terminal fields of different spinal nerves occurs due to intermingling of spinal nerves within the
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lumbosacral plexus. The degree to which they are mixed shows considerable individual variablility. This variability is referred to as prefixation or postfixation of the lumbosacral plexus. The peripheral distribution of spinal nerves differs according to whether the lumbosacral plexus is prefixed or postfixed (17, 20, 21). In the prefixed class L5 but not S2 contributes to the plexus and in the postfixed class S2 but not L5 contributes to the plexus (Fig. 1.) The amount of denervation in a muscle produced by a spared L7 nerve lesion will therefore depend on the fixation of the plexus (20, 21). For example, the tibialis anterior muscle is innervated by fibers from the spared L7 in cats with a prefixed plexus but is not innervated by L7 in cats with a postfixed plexus (21). Therefore, in spared nerve preparations, tibialis anterior will be completely denervated in postfixed animals but at least partially innervated in prefixed animals. Thus, it was possible to compare the results of partial denervation of a muscle in some animals with complete denervation of the same muscle in other animals. Differences in reinnervation and in reflex recovery between partially and completely denervated muscle were studied. Furthermore we attempted to determine if differences in reflex recovery from the lesion could be related to sprouting elicited from the spared L7 spinal nerve. MATERIALS
AND
METHODS
Twenty-two adult cats of either sex were used. Surgery was carried out under sterile conditions using Nembutal anesthesia. The spinal nerves on PREFIXED
PLEXUS
POSTFIXED
PLEXUS
L5 L6 L7 Sl s2
FIG. 1. Diagram showing the two classes of fixation of the lumbosacral plexus and the site of transection of the spinal nerves which contribute to the plexus. L5, L6, Sl, and S2 were cut and L7 was spared. In the prefixed class, L5-Sl contribute to the plexus but S2 does not. In the postfixed class, L6-S2 contribute to the plexus whereas L5 does not. In the prefixed condition the tibialis anterior muscle is innervated by L7 and in the postfixed condition it is not.
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AND REFLEX
735
RECOVERY
one side of the cord were visualized and L5 and L6 spinal nerves were cut after enlarging the intervertebral foramina; Sl and S2 were cut inside the vertebral canal. All these spinal nerves were transected distal to the dorsal root ganglia. The L7 spinal nerve was always spared. Five tendon reflexes in the cat hind limb were examined on the control and experimental sides daily for 4 postoperative weeks (Table 1). The reflexes in the experimental limbs were compared with those on the control side and graded as absent (-), weak ( +- ), strong (+), and/or clonic (c). Reflexes in the experimental limb which were indistinquishable from those on the control side were classified as normal. Reflexes and locomotion were also filmed. Five of the animals were anesthetized (two at 3 days, and one each at 7, 11, and 13 days postoperative) and either tibialis anterior or soleus and the toe flexor muscles on both legs were exposed and injected with 0.15% methylene blue in physiologic saline such that the entire muscle was stained dark blue (approximately 50 ml). Ten minutes after the injection the muscles were removed, cut in strips, placed on moist gauze, and oxygenated 1 h, at 1 liter/min. Each strip was exposed to the oxygen for 0.5 h per side. The muscle was then fixed 24 h in 8% ammonium molybdate at 4°C washed, cut into thin pieces, and squashed between two slides (4). The entire muscle was dehydrated and coverslipped and examined using a light microscope. The slides were projected and the stained nerves, arborizations, and motor end-plates were drawn with the aid of a drawing tube or were photographed. The animals were perfused through the heart with a 0.5% glutaraldehyde and 4.0% paraformaldehyde mixture. Survival periods are TABLE
1
Tendon Reflexes in Cats’ Hind Limbs Tendon reflex
Tap
Muscle(s)
Response
Hamstrings jerk Lateral (LHj) Medial (MHj)
Tendon of hamstring muscles
Biceps femoris and semitendenosus
Slight flexion of lower leg
Knee jerk (Kj)
Patellar tendon
Rectus femoris
Extension of lower leg
Ankle jerk (Aj)
Achilles tendon
Gastrocnemious and soleus
Extension of foot
Tibialis anterior jerk (TAj)
Tibialis anterior tendon
Tibialis anterior
Flexion of foot
Toe jerk (Tj)
Several tendons on plantar surface of toes
Several toe flexors
Flexion of toes
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PRENDERGAST,
shown in Table plexus, and the exposed, drawn, the lesions and
MURRAY,
AND
GOLDBERGER
2. The spinal cord, dorsal and ventral roots, lumbosacral peripheral nerves to the muscles in both hind limbs were and photographed in situ and then dissected out to confirm to determine the fixation of the lumbosacral plexus. RESULTS Behavior
Normal. Table 1 shows the occurrence of tendon reflexes in normal cat hind limbs. Tap of the tendon results in a brief, single response of leg, foot, or toes followed by return to the resting position. Normally the tendon reflexes are bilaterally symmetrical in strength. Spared Nerve. Acute period (I to 2 days postoperative). On recovery from anesthesia, the tendon reflexes in the hind limb contralateral to the lesion appeared to be normal In the partially denervated hind limb of all cats some reflexes were lost and other reflexes were weak compared with reflexes on the contralateral side. The animals were assigned to two groups on the basis of the status of their tendon reflexes. These animals could be assigned to the same two groups based on the fixation of their lumbosacral plexus, as determined by dissection after killing (Table 3). (i) Prefixed class (N = 9): The tibialis anterior and toe jerks were always present, the ankle, lateral hamstrings and knee jerk were always abolished, and the medial hamstrings reflex was present in 2 cats but absent in 7 cats. (ii) Postfixed class (N = 13): The ankle jerk was present, and the tibialis anterior and knee jerks were lost. The toe jerk was present in 8, the medial hamstrings jerk in 5, and the lateral hamstrings jerk in 11 of the cats. Early recovery period (3 to 7 days). (i) Pre$xed class (N = 9): The responses which were present in the immediate postoperative period became progressively stronger. In addition, tapping the region of the patella, presumably including stimulation of the periostium, produced internal or external rotation of the leg and protraction of the hip in 8 of these cats. TABLE
2
Times of Killing after Lesion and Numbers of Cats in Each Class of Fixation of the Lumbosacral Plexus Acute (1 to 2 days)
Early (3 to 7 days)
Chronic (8 to 19 days)
Prefixed
0
1
8
Postfixed
1
4
8
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AND REFLEX
TABLE
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3
The Pattern of Reflexes Which Were Elicited during the Recovery Period in one Cat Whose Lumbosacral Plexus Was Prefixed and Another Whose Plexus Was Postfixed Prefixed”
Postfixed
Acute (1 to2 days)
Early (3 to I days)
Chronic (8 to 17 days)
Acute (1 to2 days)
Early (3 to 7 days)
Chronic (8 to 19 days)
LHj MHj
(-) (-)
(-) C-J
(+) (+I
(-) (-)
(-) (-)
(-) (+)
Kj
(-)
(-)
(+)
C-J
(-)
C--J
TAj
(+-)
(+I
(+)
(-)
C-1
(+)
Aj
C-1
(-)
(*)
(k)
t+. c)
(+I
Tj
(*I
(+I
(+I
(*I
(+)
(+I
Tendon reflex’
” (-), and Hj cat the h For
Absent; (+), strong; (k), weak; (c), clonic. Notice in the prefixed cat that the Aj return, the TAj and Tj become stronger, and the Kj never recovers. In the postfixed TAj and LHj recover, Aj and Tj become stronger, and the Kj never returns. abbreviations see Table I.
These responses, which are not normally seen, are presumed not to be tendon reflexes because tap of the patella tendon alone produced no response. (ii) Postjixed class (N = 12): The ankle jerk, which was weak initially became stronger and then clonic in 3 cats. In 10 of the 12 cats which survived into this period, tap of the patellar region produced the same rotation and protraction responses as in the prefixed class. Chronic period (8 to 19 days). (i) Prefixed class (N = 8): The ankle jerk returned in 3 cats in which it had been absent. The lateral hamstrings jerk returned in 2 cats in which it had been absent. In 3 cats the tibialis anterior jerk, which had been weak, became strong and then clonic. (ii) Postjixed class (N = 8): Of the 8 cats in this group, the lost tibialis anterior returned in 3 and the lost toe jerks returned in three. Summary of Tendon Reflexes
On the day after surgery the pattern of reflex loss was correlated with the fixation of the lumbosacral plexus, In the prefixed class the ankle, knee, and lateral hamstrings jerks were lost, and the remaining reflexes were weakened. By 7 days all weakened reflexes had become stronger. During the following 12 days the abolished ankle and lateral hamstrings jerk re-
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turned. In the postfixed class in the acute period the tibialis anterior and knee jerks were absent in all cats. In the chronic period the abolished toe and tibialis jerks returned in some cats. The pattern of reflexes which were spared were therefore different for the two classes but in both classes (i) the weakened reflexes became stronger; some even became clonic, and (ii) some of the lost reflexes returned within 3 weeks after the lesion. Anatomy
The following description of morphological changes treats only the tibialis anterior in detail. The changes occurring in tibialis were, however, representative of those occurring in soleus and the toe flexors. Normal (unoperated side, N = 5). The intramuscular nerves coursed across and in between the muscle fibers. The large intramuscular nerves contained many axons, were relatively straight, and branched often. The branches always contained at least five axons. The axons also had a characteristic branching pattern and finally splayed out in a terminal arborization. Usually a single, but occasionally two motor end-plates were seen to arise from each preterminal axon. An example of the characteristics of the normal intramuscular nerves, the axons within them, and the motor end-plates is shown in Fig. 2. Spared nerve (N = 5). Early postoperative periods [(3 and 7 days) (N = 3)]: The morphology*of the tibialis anterior intramuscular nerve on the lesion side differed from normal and also differed between pre- or postfixed animals. In both classes, degenerating axons were present and these were swollen and beaded as they coursed across the muscle (Figs. 3, 4). The motor end-plates on the degenerating terminal axons were swollen and clumped (Fig. 4B, C). Prefixed class (N = 1): The intramuscular nerve was characterized by the presence of some normal appearing axons coursing together with the degenerating axons (Fig. 3). Postfixed class (N = 2): All axons and motor end-plates (Fig. 4) in the tibialis anterior muscle were degenerating. Only a few degenerating motor end-plates were still detectable by 7 days after surgery. Chronic period [ ( 11 and 13 days) (N = 2) 1: In both classes there was some degenerating debris remaining in the muscle. The characteristics of the intramuscular nerve again differed betweeen preand postfixed animals. Prefixed class (N = 1): The pattern of branching of the axons in the intramuscular nerve was identical to that of the control side. Frequently, however, as many as six or seven end-plates branched from a single axon, a condition never seen normally. (Figs. 5, 6). PostJxed class (N = 1): There were a few nondegenerating axons in each intramuscular nerve bundle. A division of the intramuscular bundle often resulted in a single fiber coursing across the section of muscle without branch-
FIG. 2. A-photomicrograph showing an intramuscular nerve and one of its branches stained with methylene blue in a normal cat tibialis anterior muscle preparation, The arrow indicates the characteristic branching pattern of the nerve fibers within the intramuscular nerve. X250. B-photomicrograph showing several motor end-plates (stars) arising from the fibers of a normal terminal arborization. Some motor end-plates are out of the plane of focus (double stars) due to the thickness of the squash. X250. Methylene blue stain.
2 \”
740
PRENDERGAST,
MURRAY,
FIG. 3. Photomicrograph of tibialis anterior class, during the acute postoperative period (3 degenerative nerve fibers (arrows) as well as within the intramuscular nerve of the muscle.
AND
GOLDBERGER
muscle and nerve from a cat of the prefixed days postoperative). Note the swollen, beaded normal nerve fibers (stars) coursing together X325. Methylene blue stain.
PERIPHERAL
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741
RECOVERY
‘6’ A
FIG. 4. Photomicrographs and projection drawing of tibialis anterior muscle and nerve in a postfixed cat. The tibialis anterior muscle contains only swollen, beaded degenerating nerve fibers in the intramuscular nerve and swollen degenerating motor end-plates. 7 days postoperative. A-a large intramuscular nerve, in which all nerve fibers are degenerating. X150. Methylene blue stain. B-three degenerating nerve fibers, one of which terminates in a degenerating motor end-plate (DMEP). X500. Methylene blue stain. C.-Projection drawing of the degenerating nerve fibers and motor end-plate (DMEP) shown in B through several planes of focus.
742
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MURRAY,
AND GOLDBERGER
FIG. 5. Photomicrograph shows four terminal nerve fibers (a-d), from one (c) of which three motor end-plates emerge. X500. The star indicates the motor end-plate most nearly in focus. Prefixed cat, 1 I days postoperative, methylene blue stain.
ing. Along the course of the axons, knot-like regions were encountered (Fig. 7). These were never seen in normal muscle or in partially denervated tibialis anterior muscle from cats in the prefixed class. It was not possible to distinguish whether these knot-like thickenings were a point of repeated branching or simply a tangle of nerves. Commonly, however, there were more axons on one side of the thickening. Several terminal axons extended
\
-h
MEP
FIG. 6. A terminal nerve fiber: Three-dimensional, photographic reconstruction through several planes of focus. The negatives of photomicrographs taken at different focal planes were stacked upon one another and photographed to produce the positive. This procedure preserved in detail the terminal nerve fiber and the motor end-plates (MEP) which emerge from it. The reconstruction shows a single nerve fiber from which six motor end-plates emerge. X1 12.5. Prefixed cat, 11 days postoperative.
PERIPHERAL
SPROUTING
AND
REFLEX
RECOVERY
743
from the thickening (Fig. 8) each of which ended in a single, normalappearing motor end-plate. Summary
of Anatomy
Morphological changes in the nerve to tibialis anterior muscle varied as a function of postoperative time and the class of fixation to which the lumbosacral plexus belonged. In the prefixed class in the early postoperative stages the nerve contained bundles of normal and degenerating axons. Each normal axon ended in a single motor end-plate. By 13 days there were several motor end-plates from each preterminal axon but the branching pattern of the axons in the intramuscular nerve appeared normal. In the postfixed class, in the early stages only degenerating axons and motor endplates were seen. In the chronic postoperative periods, however, a few axons were seen in the nerve and these contained knot-like thickenings which appeared to be branch points because the number of axons on one side of the thickening always exceeded the number on the other side. Some of these axons ended in a single, normal appearing motor-end plate. General Summary
(Table 4)
In the prefixed class at 3 days after surgery the reflex was weak. The muscle was presumed to be partially denervated because normal and degenerating axons were seen in the same bundle. One, and occasionally two motor end-plates extended from preterminal axons. Multiple motor endplates were seen to arise from single axons. In the postfixed class acutely, the tibialis anterior jerk was absent; no axons were seen in the muscle except those that were degenerating. In the chronic period, in those animals in which the reflex returned some axons were seen in the muscle. These axons contained knot-like structures from which additional axons extended. Some of these axons terminated in single, apparently normal, motor endplates. DISCUSSION Our results show that after transecting L5, L6 and Sl, S2 (sparing L7) spinal nerves, (i) two different distributions of tendon reflexes are seen in the hind limb ipsilateral to the lesion, (ii) some of the reflexes which were weak acutely became stronger and even clonic later, and (iii) some of the reflexes which were abolished for as long as 2 weeks returned. The two reflex patterns seen acutely in the hind limb depended on the class of fixation of the lumbosacral plexus. In both classes, the reflexes which re-
144
PRENDERGAST,
MURRAY,
AND GOLDBERGER
i’-j-L Experimental
C FIG. 7. Branching of an intramuscular nerve. A.--Camera lucida drawing of a control. B.Three examples of camera lucida drawings of branching of the intramuscular nerves containing knot-like regions described under Results. Postfixed cat, 13 days postoperative. C and D-C is a drawing of branching of a nerve containing two knot-like regions and D is the photomicrograph (X350) from which the drawing (C) was made. Methylene blue stain. The arrow is pointing to a distal branch and shows the orientation. 1 and 2 refer to two knot-like regions and P indicates the presumptive parent axon. Postfixed cat, 13 days postoperative.
mained after the lesion became stronger beginning on the third postoperative day. When the muscle in which weak reflexes could be elicited in the acute period was examined histologically, some normal axons were
PERIPHERAL
SPROUTING
AND
REFLEX
RECOVERY
745
always present in the intramuscular nerve. The terminal arborization of the intramuscular nerve was characterized by one, and very occasionally two, motor end-plates per axon. When examined in the chronic period, an increased number of motor end-plates was seen emerging from a single axon. The branching pattern of the intramuscular nerves and the axons within it were normal. The increased number of motor end-plates per axon indicated that sprouting had occurred and had reinnervated the partially denervated muscle. The sprouting apparently occurred early in the first postoperative week concomitant with an increase in strength of the reflex. The axonal sprouts presumably arose from axons which normally innervated the partially denervated muscle. We therefore call this “homonymous” sprouting. It is comparable to sprouting which has classically been called collateral sprouting (8) and has been correlated with several aspects of recovery of function after peripheral nervous system lesions, (2, 7, 12, 14, 16, 18, 25-27). In the tibialis anterior in postfixed animals, no tendon reflex could be elicited in the acute stage. When this muscle was examined in the early, areflexic postoperative stage, only degenerating axons and degenerating motor end-plates were encountered. This is consistent with the observation that in cats with postfixed lumbosacral plexi, electrical stimulation of L7 does not elicit contraction of tibialis anterior (2 1). These observations therefore indicate that in the early stages after surgery the tibialis anterior had no innervation from the spared L7 in postfixed animals. In some animals, the lost reflex returned in the chronic periods. When the tibialis anterior was examined histologically after this reflex recovery, a small number of axons were found. These axons were abnormal in containing knot-like regions from which branches appeared to emanate. Some of these branches coursed singly across the tissue without further branching and others penetrated various distances through the muscle singly or in small groups and terminated with single, normal-appearing motor end-plates. Similar abnormal knot-like regions were described in the course of an examination of muscle in patients whose only other distinction was that of mental illness (5). We interpret our observations to mean that axons had sprouted into the tibialis anterior to reinnervate this denervated muscle. Because in such postfixed animals, all nerves to tibialis anterior had degenerated during the areflexic period, the sprouted axons presumably arose from a nerve innervating another muscle. We propose to call this “heteronymous” sprouting. This differs from the classical description of collateral sprouting in that it apparently does not depend on overlap, i.e., the transected and sprouting nerves do not innervate the same muscle preoperatively. Innervation by sprouting or regeneration by a foreign nerve into a zone previously not
746
PRENDERGAST,
MURRAY, TABLE
AND GOLDBERGER 4
Time Course and Character of the Changes in the Reflex and in the Nerves of the Tibialis Anterior Muscle after Transection of Ls, L,,, S,, and S2 (L, Spared) Early (3 to 7 days)
Chronic (8 to 19 days)
Reflex
Anatomy of nerves
Reflex
Anatomy of nerves
Prefixed
Weak
Degenerating and normal
Strong
(i) Several motor end-plates per terminal fiber. (ii) Normal branching of intramuscular nerve and nerve fibers.
Postfixed
Absent
All degenerating
Weak
(i) Few fibers in intramuscular nerve. (ii) Gnarled regions along nerve fibers. (iii) Fibers from gnarled zone course futher or terminate with single motor endplate.
supplied by that nerve has been described in the peripheral nervous system (1, 3, 10, 13, 18, 22-24, 28). Thus two different patterns of reflex recovery can be correlated with two different forms of sprouting. Some of the reflexes which were abolished by surgery never returned. One of these was the knee jerk. Tap of the region of the patella, e.g., skin and presumably periostium, however, resulted in abnormal motor responses at the hip beginning at 3 days after surgery. These responses could have resulted from the unmasking of latent or inhibited reflex pathways when the knee jerk was abolished and this process might require 2 or 3 days after the surgery before the reflexes could be elicited. On the other hand, the time course of their appearance suggested that they also might result from axonal sprouting. The present experiment demonstrates that recovery of a reflex, i.e., a centrally integrated function, which occurs in less than 1 month after cutting spinal nerve in cat, must be due, at least in part, to sprouting of axons which survive the lesion. Early “homonymous” sprouting or later “heteronymous” sprouting always resulted in adaptive recovery. For example ( 1l), when the tibialis anterior jerk was absent, the foot always slipped backward during stepping, but when this reflex returned no slipping during stepping was seen.
PERIPHERAL
SPROUTING
AND REFLEX
RECOVERY
747
B FIG. 8. Terminal nerve fibers as they branch from a gnarled region of an intramuscular nerve. In the photomicrograph (A) and the reconstruction (B) the fibers labeled (a, b, c) and motor end-plates labeled (1, 2, 3) are the same. A-photomicrograph showing two nerves. From one (arrow), three branches arise (a, b, c) and terminate in motor end-plates (1, 2, 3). Notice that parts of the terminal nerves and the motor end-plates are out of the plane of focus due to the thickness of the squash. Postfixed cat, 13 days postoperative, methylene blue stain. B.-this three-dimensional photographic reconstruction was made through several planes of focus by the same method as Fig. 6 and shows the three branches (a, b, c) and the motor endplates (1, 2, 3) which arise from them. The negative for photograph (A) above was 1 of 10 used to make this reconstruction. Notice that the terminal fibers and the motor end-plates (MEP) can be seen in more detail in the reconstruction than in the photomicrograph (A).
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PRENDERGAST,
MURRAY,
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