Normal Innervation Pattern Research Articles

The effects of black widow spider venom on the innervation of muscles paralysed by botulinum toxin.

Botulinum toxin (BoTx) was injected into the muscles of one leg in mice causing local paralysis. Black widow spider venom (b.w.s.v.) was then injected into the paralysed muscles 3 or 15 d later. In both groups b.w.s.v. destroyed the nerve terminals poisoned by BoTx. In the 15 d group axonal sprouts, which had former due to the block of neuromuscular transmission by BoTx, were also destroyed. Within a few days the motor nerve terminal regenerated and the muscles recovered from paralysis at a faster rate than after BoTx alone. Recovery seemed to begin earlier in muscles where axonal sprouting was already advanced when b.w.s.v. was injected. The normal pattern of innervation was re-established in both groups, which was in marked contrast with muscles after BoTx alone where numerous sprouts and many ectopic end-plates had formed.

Supraoptic nucleus of the Brattleboro rat has an altered afferent noradrenergic input.

The distribution of fluorescent varicosities in the supraoptic nucleus of Brattleboro rats was compared to that in normal rats. The Brattleboro rat, which is characterized by a genetic absence of vasopressin, had fewer fluorescent varicosities in apposition to the vasopressin-deficient perikarya. The oxytocin-producing neurons in the same nucleus were hyperinnervated. These data suggest that the target neuron peptide (vasopressin) is necessary for the maintenance of normal noradrenergic innervation patterns.

The neuromuscular junction of the mouse after black widow spider venom.

1. A sublethal quantity of black widow spider venom was injected into the calf muscles of mice. After 30 min to 6 weeks soleus muscles were examined by light and electron microscopy and by electrophysiological techniques. 2. Within 30 min motor nerve terminals were swollen and depleted for synaptic vesicles and by 6 h were disrupted and engulfed by Schwann cells. By 24 h every end-plate examined was denervated. Some preterminal myelinated axons also showed degenerative changes. 3. Re-innervation was first seen at 2 days. By 3 days axon terminals were present at most end-plates and by 8 days their morphology was nearly normal. The normal pattern of innervation of the muscle was re-established in that axons re-innervated their original end-plates and very few ultraterminal axonal sprouts were found. 4. Physiological study showed complete failure of transmission and absence of miniature end-plate potentials (m.e.p.p.s) and end-plate potentials (e.p.p.s) until day 3, when muscles responded weakly to indirect stimulation and m.e.p.p.s were recorded at 30% and e.p.p.s at 40% of fibres. The mean quantal content of e.p.p.s was low and there was rapid fatigue on repetitive stimulation. Extrajunctional sensitivity to acetylcholine developed within 1 day, was maximal at 3 days and declined to normal at 12-14 days. 5. The proportion of fibres at which m.e.p.p.s and e.p.p.s were recorded returned to normal by day 6 and mean quantal content was normal by day 9. 6. These findings show that the re-innervation of original end-plates is of importance in facilitating the rapid return of transmission to normal levels and limiting the extent of axonal growth.

The innervation of dorsoventrally reversed chick wings: evidence that motor axons do not actively seek out their appropriate targets

In normal chick embryos the extensor (dorsal) muscles are innervated by motoneurones lying laterally in the motor horn, while flexor muscles are supplied by more medially placed motoneurones. After reversal of the dorsoventral axis of the forelimb prior to innervation in most cases the opposite pattern is found, the extensors innervated by medial and flexors by lateral motor neurones. In a minority of cases the normal innervation pattern is obtained. Three hypotheses are discussed, two involving specific target affinity between motor axon and target and one involving passive deployment of axons to targets. We conclude that our results favour the latter hypothesis but that we cannot exclude a short-range specific signal.

A quantitative assessment of muscle spindle formation in reinnervated and non-reinnervated grafts of the rat extensor digitorum longus muscle

Free grafting of the rat extensor digitorum longus muscle disrupts the neurovascular supply and results in degeneration and subsequent regeneration of both muscle and nerve. Reinnervation of one group of grafts was prevented by severing and ligating the sciatic nerve. Thirty days after surgery, the number of muscle spindles in each graft was determined by examination of serial sections at the light-microscopic level. Non-operated control muscles contained 37.86 ± 3.02 ( s.d.) spindles per muscle. Means for the reinnervated and non-reinnervated grafts were 15.63 ± 3.89 and 18.38 ± 5.15 respectively. At the time of surgery, all of these grafts had been treated with 0.75% bupivacaine, a local anesthetic which, combined with the grafting technique, ensures degeneration of all muscle fibers. An additional group of grafts was not subjected to this treatment and contained a mean of 27.40 ± 2.19 spindles. The embryogenesis of muscle spindles is dependent upon interaction between sensory nerves and developing myotubes. The conditions for the formation of these stretch receptors in regenerating muscle have been less clear. Their formation in non-reinnervated grafts demonstrates that this process, unlike spindle development in the embryo, does not require the presence of nerves. When reinnervation of a graft is allowed, large numbers of nerve fibers have been observed in muscle spindles. It is not yet known whether or not the normal pattern of innervation, essential for proper function of the spindle, is restored.

The production and storage of Nerve Growth Factor in vivo by tissues of the mouse, rat, guinea pig, hamster, and gerbil

The Nerve Growth Factor (NGF) content in vivo of tissues from the mouse and rat at various stages of development from 3 days embryonic gestation to the attainment of full maturity has been determined using the standard biological assay. A less extensive survey has also been made of tissues from the guinea pig, hamster, and gerbil. With the exception of the well-documented high levels of NGF in the mouse submaxillary glands, none of the organs examined contained detectable NGF. These results, which are consistent with those previously reported using the biological assay, stand in contrast to the high levels of NGF detected in virtually all tissues by some published radioimmunoassays. It is likely that the discrepancies are due to the use in the radioimmunoassays of antisera containing antibodies to proteins other than NGF, and to the inability of one-site radioimmunoassays to distinguish between the presence of NGF and that of agents capable of binding NGF. The apparent lack of widespread NGF production in vivo contrasts with the ability of many tissues to synthesize the protein in vitro. This may imply that physiologically significant levels of NGF are below the limits of sensitivity of the assay systems presently available, that NGF synthesis in vivo occurs only during a very restricted period of development, or that the presence of a normal innervation pattern influences NGF production.

Organization of brainstem noradrenaline hyperinnervation following neonatal 6-hydroxydopamine treatment in rat.

The organization and origin of NA axonal sprouting in individual brainstem nuclei was examined by fluorescence histochemistry and a radioenzymatic assay for noradrenaline (NA) in adult rats which were administered 6-hydroxydopamine (6-OHDA) as neonates. Significant 2–6 fold increases in NA content were measured in primary sensory nuclei, associational nuclei, somatic motor and visceral cranial nerve nuclei, reticular formation, raphe nuclei, and the inferior olivary complex. Fluorescence histochemical analysis reveals a major increase in the number of fluorescent fibers in most areas of the brainstem after 6-OHDA treatment. The normal pattern of innervation and axon morphology of each nucleus is retained after drug treatment. Bilateral locus coeruleus (LC) lesions in treated animals results in the denervation of only those nuclei which normally receive LC innervation (Levitt and Moore, 1979). The increased number of NA-containing axons in nuclei innervated by the lateral tegmental cell groups (LT) remains intact after LC ablation. The present results indicate that the brainstem NA sprouting following neonatal 6-OHDA administration occurs in a highly specified manner. The brainstem projections of both the LC and LT neuron groups exhibit marked axonal growth which is restricted to their specific brainstem terminal fields and probably occurs in response to the denervation of their respective, more rostral target areas.

Regeneration of normal terminal innervation patterns by central noradrenergic neurons after 5,7-dihydroxytryptamine-induced axotomy in the adult rat

The regeneration of central adrenergic axons has been followed between 5 days and 18 months after 5,7-dihydroxytryptamine (5,7-DHT)-induced axotomy in the adult rat, using fluorescence histochemistry in combination with noradrenaline (NA) determinations and [ 3H]NA uptake measurements. The axonal and terminal degeneration caused by the 5,7-DHT treatment (150 μg intraventricularly) was, by 1–2 weeks after injection, accompanied by a 70% reduction of NA in the forebrain and 30% reduction in the brain stem, and by 43–85% reductions in the [ 3H]NA uptake capacity in various regions of the brain and spinal cord. Signs of sprouting of the drug-lesioned axons were evident along the terminal axon segments at 5 days after treatment. The sprouts increased rapidly in length and number during the subsequent weeks and by 2–6 months after injection new NA terminal systems of relatively normal density and distribution had been re-established in many initially denervated regions. In parallel there was a recovery of endogenous NA and [ 3H]NA uptake to the pre-injection levels in the brain, and to levels 50–75% of normal in the cervical and thoracic spinal cord. Four successive phases of the regeneration process are distinguished: (1) primary sprouting from the lesioned NA axon stumps, occurring within the first week after treatment; (2) seemingly random growth and proliferation of the newly formed sprouts during the second and third weeks; (3) directed, forward growth of some of the sprouts leading to a partial restoration of the original fibre paths, branching patterns and terminal networks within 3–6 months; (4) a concomitant removal of at least part of the abnormally directed sprouts. Although the original fibre architecture was quite accurately restored in many areas the regeneration was not always correct. Hyper-innervation patterns and abnormal terminal arrangements were often formed, and in the spinal cord the down-growth of the regenerating axons occurred predominantly along a route that is inconspicuous in the normal rat. It is concluded that at least certain types of central neurons regenerate very efficiently provided the conditions are favourable, and that under such conditions axonal regeneration in the mammalian CNS is subjected to regulatory mechanisms that can be very precise. The results provide evidence that the adult mammalian CNS possesses mechanisms for axonal guidance which allow the accurate regeneration of lesioned axonal tracts and branching patterns, as well as mechanisms of recognition making possible the re-establishment of the terminal connections.

Ultrastructure of the muscle spindle in dystrophia myotonica. II. The sensory and motor nerve terminals.

The normal pattern of innervation was lost in the four dystrophia myotonica muscle spindles examined. There had been a proliferation of both sensory and motor nerve terminals, and many endings showed varying degrees of structural abnormality. The changes seen in the sensory nerve endings were an incomplete contact relationship between the terminal and the muscle fibre, the association of Schwann cell processes with the nerve ending, abnormal shape of the terminal, multiple sensory endings, and changes in the ultrastructural organization of the terminal axoplasm. Remnants of degenerated sensory terminals, and isolated sensory endings were also seen. Motor nerve terminals varied in size and shape. In general, they contained a normal complement of vesicles and mitochondria. Junctional folding was often absent or simplified, and sarcoplasmic specialization was, at the most, rudimentary. The relationship between these changes and intrafusal muscle fibre fragmentation is discussed.

Monoamine neuron innervation of the hippocampal formation: Alteration by neonatal irradiation

Previous studies showed that postnatal irradiation of the hippocampal formation in the rat results in a marked reduction in the granule cell population of the area dentata with an attendant disruption of its normal cytoarchitecture. The present study was carried out to determine the effect of postnatal hippocampal irradiation on the pattern of hippocampal innervation by locus ceruleus norepinephrine neuron axons and midbrain raphe serotonin neuron axons. Irradiated animals exhibit a normal innervation of Ammon's horn by both norepinephrine and serotonin neuron axons. In the area dentata, however, the normal pattern of innervation is markedly altered, with both an apparent decrease in total innervation and a redistribution of the innervation occurring. These effects are interpreted as representing a failure of development of the growth-promoting factors that determine the normal innervation pattern.

Regeneration of fungiform taste buds: temporal and spatial characteristics.

The gross morphology of the tongue of the Mongolian gerbil Meriones unguiculatus), the location of papillae and taste buds, and the normal innervation pattern of the tongue and taste buds were determined. The chorda tympani nerve was interrupted to produce degeneration of fungiform taste buds. Regenerating chorda tympani axons followed the original nerve pathways in the tongue en route to the fungiform papillae in the epithelium where they initiated the regeneration of taste buds. The spatial distribution of reinnervated fungiform papillae and reformed taste buds was examined 7 to 19 days following surgery. Beginning at eight days following chorda tympani interruption there was a progressive increase, first, in the proportion of fungiform papillae that were reinnervated, and later in the number of reformed taste buds. On the basis of these measures it was concluded that a taste bud is reformed one to two days after reinnervation of its papilla. From the time course of reinnervation of the fungiform papillae it was calculated that some fibers regenerated at rates in excess of 2 mm/day. Regeneration was precise and systematic. The regenerating chorda tympani fibers accurately returned to the fungiform papillae; they did not follow the pathways of lingual nerve axons. In the initial stages of recovery both reinnervated papillae and reformed taste buds were preferentially located toward the front of the tongue; the reinnervation of posterior fungiform papillae was delayed.

Growth in culture of the peripheral axons of the spiral neurons in response to displacement of the receptors.

Observations were made on the growth of the peripheral axons of the spiral neurons in injured explants of the cochlea of the mouse. Opening of the cochlear duct during explantation usually results in a displacement of the hair cells into the outgrowth zone if the basilar membrane of the excised organ of Corti faces the substrate. Spiral neurons respond with vigorous growth to the stress created by the displacement of their receptors. Two different growth reactions occur in succession: 1) growth by elongation of the synaptically engaged fibre 2) free growth. The response of the neuron to an incipient displacement of the hair cells is an elongation of the fibre while its contacts with the receptors are, at least partly, preserved. Most or all fibres that are connected with the displaced cells elongate. The growth is well organized, limited to the region of the displaced hair cells, and restricted in length by the position of the receptors. It is inferred that tension on the synaptically engaged fibre may be a stimulus for its growth and that the growth ceases when the tension is relieved. Continuous stress eventually leads to a rupture of the fibre. The break usually occurs near or at the terminals, leaving the terminals attached to the hair cells. The proximal end of the fibre--now free--starts to grow. The growth is independent of the receptors, unrestricted in length, uncontrolled in amount, and continues for at least three weeks after explantation. The path of the fibre is tortuous, and collaterals are often emitted. Free growth proceeds at a markedly faster rate than growth by elongation. Maintenance of the synapse and growth of the fibre seem to be mutually dependent events. A peripheral axon of the spiral neuron can sustain in culture a normal innervation pattern, without any obvious signs of abnormal growth, if its connections with the hair cells are undisturbed; it elongates in an organized manner, if these connections come under stress; it responds with an uninhibited growth if the connections are broken.

Growth patterns of the peripheral axons of the spiral neurons in culture

Segments of the organ of Corti with its innervation preserved were excised from the newborn mouse and studied in culture. The following growth patterns were observed: (a) maintenance of the normal innervation pattern and growth along normal lines if the hair cells remain in their usual location, (2) growth by elongation of the fiber if the hair cells migrate into the out‐growth zone and if the fiber remains in contact with the receptor, and (3) free growth of the fiber if its connections with the hair cells are broken. Growth by elongation is confined to the area of hair cell displacement. The growth is well organized and the length of the fiber compensates strictly for the distance the respective hair cell has migrated. Free growth proceeds at a faster rate then does growth by elongation, is independent of the receptors, is unrestricted in length, and uncontrolled in amount. The growth is disorganized and luxuriant for at least several weeks after explanation. [Work supported by NIH grants NS08626 and NS12732.]

A search for the factors responsible for absence of recovery of the normal vascular response to oxytocin following sympathetic nerve injury.

In dogs following crush injury to the lumbar sympathetic trunk, reflex vasoconstriction reappears in 4-6 months but the normal vasodilator response to oxytocin does not return even 12 months after crush. Histochemical examination of the walls of the blood vessels shows that division or crush of the lumbar sympathetic trunk or removal of terminal ganglia leads to decentralization, not denervation of the blood vessels. True denervation follows division or crush of the sciatic and femoral nerves. Following recovery from sciatic or femoral crush the pattern of peripheral innervation appears histochemically normal. However, there is no return of the normal vasodilator response to oxytocin. It is concluded that a normal response to oxytocin does not return even after long-term recovery from sympathetic injury, nor does its effect depend on a normal pattern of peripheral adrenergic innervation, but on an unknown more central activity of the sympathetic nervous system.

Studies on the development of the chick optic tectum. IV. An autoradiographic study of the development of retino-tectal connections

The normal pattern of innervation of the optic tectum has been studied autoradiographically in a series of chick embryos which were injected intraocularly with [ 3H]proline at intervals between the 6th and 21st days of incubation. From the distribution of the radioactively labeled proteins transported in the rapid phase of axonal flow, it is evident that retinal fibers first enter the tectum late on the 6th day of incubation and then spread across its surface from its rostrolateral aspect to its caudo-dorso-medial pole during the ensuing 6 days. At the 9th day, when the fibers have grown across the surface of the rostral half of the tectum, there is no indication that the terminal portions of the axons have left the stratum opticum to enter the outer layers of the stratum griseum et fibrosum superficiale. The first suggestion of such an invasion of the straum griseum et fibrosum superficiale is found at day 10, when labeled fibers can be seen in this stratum, over a restricted, oval, area near the center of the tectum. Over the course of the next 2 days the region of the stratum griseum et fibrosum superficiale occupied by retinal fibers expands, more-or-less concentrically, until by day 14 some labeled fibers are seen in the outer part of the stratum, throughout the tectum. Concurrent with the ingrowth of optic nerve fibers the final cytoarchitectonic differentiation of the outer layers of the tectum occurs so that by day 18 the autoradiographs show a pattern of labeling of layers a–f of the stratum griseum et fibrosum superficiale similar to that seen in mature, post-hatched chicks. Since the region of the tectum which is first innervated by retinal fibers corresponds to the projection field of the region around the upper end of the choroid fissure where the first ganglion cells are generated, it would appear that the axons of the first-formed ganglion cells grow over the rostral surface of the tectum before establishing the first retino-tectal synapses near the central portion of the tectum. Subsequently, as the ganglion cell population grows concentrically from around the area centralis, there is a parallel expansion of the region in which retino-tectal synapses are being formed in the tectum.

Implant-induced accessory limbs in urodeles: fresh, frozen, and boiled tissues.

AbstractImplants were made into forelimbs of Triturus viridescens using fresh, frozen and boiled kidney and liver of T. viridescens and R. pipiens. Limbs were recovered at intervals up to 70 days post‐implantation.Kidney implants from Wisconsin R. pipiens gave twice as many extensive accessory structures as did Vermont frog kidney. Total induction percentages, however, were similar.Quantitative and qualitative parameters for implant‐induction of accessory structures were investigated. The decrease in antigenicity and increased rate of cytolysis of frozen implants resulted in increased similarity between frog and newt kidney in rate and pattern of breakdown and in rates of induction. Modification of rate and duration of the release of the stimulating factor from the implant did not result in induction by liver implants.No evidence was found for any increase in innervation prior to or coincident with blastema formation. Implantation and implant cytolysis may cause hypersensitivity of limb tissues to the normal innervation pattern or trophic stimuli from the implant may act with those from the injured limb tissues to produce growth.The general pattern of host reaction to the implanted material was studied and described.