Dorsal Root Terminals Research Articles

Plasticity of complex terminals in lamina II in partially deafferented spinal cord: The cat spared root preparation

Projections to the dorsal horn change in adult mammals in response to complete or partial deafferentation. The number of synaptic terminals remains constant after complete lumbosacral deafferentation, indicating replacement of lost dorsal root terminals by newly formed terminals from spared intrinsic systems. The density of a spared central projection of a dorsal root is increased in dorsal horn after partial deafferentation, consistent with sprouting by the axons in the spared root. In this study, we have used electron microscopy to study morphological changes in a specific class of terminals in the dorsal horn induced by partial deafferentation. Complex terminals (CTs) in the dorsal horn originate exclusively from dorsal roots and are readily distinguished morphologically. The CTs and the postsynaptic densities (PSDs) associated with CTs were measured in lamina II at L5 and L6 in cats subjected to unilateral spared root (L6) dorsal rhizotomies and compared to CTs in the control side. Acutely following partial deafferentation, the number of CT profiles decreased. At more chronic survivals, the number of CT profiles were restored to normal levels, and both the number and the length of PSDs were increased. The changes in CTs and PSDs suggest sprouting and synaptogenesis by the spared dorsal root fibers that produce changes in the postsynaptic neuron. Spared root deafferentation thus elicits compensatory changes in presynaptic terminals of the spared root and also in their postsynaptic target neurons.

Vasoactive intestinal polypeptide in dorsal root terminals of the rat spinal cord is regulated by the axoplasmic transport in the peripheral nerve

Vasoactive intestinal polypeptide (VIP) immunoreactivity in the upper spinal dorsal horn is markedly increased after transection, crush or vinblastine treatment of the ipsilateral, segmentally related peripheral nerve. After regeneration of the peripheral nerve, VIP disappears from the upper dorsal horn. Transection-induced VIP increase is abolished by rhizotomy. It is concluded that the expression of VIP is restricted by factor(s) carried by retrograde axoplasmic transport to dorsal root ganglion cells.

Reinnervation of transplanted Pacinian corpuscles by ventral root axons: Ultrastructure of the regenerated nerve terminals

This study addresses two questions. Can mature, denervated and transplanted Pacinian corpuscles accept innervation from motor axons? If so, does the alien target influence the structural characteristics of the regenerated motor axon terminals? Pacinian corpuscles from the hind leg of young rats, together with a segment of the nerve branch through which they receive their sensory innervation, were autotransplanted to the surface of the spinal cord and the nerve stump anastomosed to the central stump of a transected lumbar ventral root. Between 4 and 5 months later the grafts were studied by electron microscopy. Ventral root axons regenerated through the endoneurial tubes of the grafted nerve to reach the corpuscles, most of which became reinnervated by one to three myelinated fibres. The fibres lost their myelin sheaths before entering the inner core, branched, and gave rise to multiple terminals in the inner core. The regenerated terminals were packed with spherical synaptic vesicles and closely resembled normal motor nerve terminals. Thus motor axons are able to reinnervate Pacinian corpuscles but the structural characteristics of the terminals are apparently not modified by the alien target tissue. This finding contrasts with previous studies, in which it was found that terminals of the central axons of large dorsal root ganglion cells, induced to reinnervate Pacinian corpuscles, displayed the structural characteristics of peripheral sensory endings rather than those of dorsal root terminals in the spinal cord.

Extracellular labeling of unmyelinated dorsal root terminals after WGA-HRP injections in spinal ganglia

Wheat germ agglutinin conjugated to horseradish peroxidase (WGA-HRP) is a widely used neuroanatomical tracer. When compared with other tracers, WGA-HRP may preferentially label unmyelinated fibers. In agreement with this hypothesis, injections of WGA-HRP in cervical and lumbar dorsal root ganglia resulted in more prominent light microscopical labeling in superficial than deep laminae of the dorsal horn. However, ultrastructural examination of these laminae reveals a paucity of terminal labeling in contrast to the abundance of extracellular tracer in the space surrounding unmyelinated fibers and their terminals, and to the widespread occurrence of transneuronal labeling. These results bear upon the mechanism of preferential labeling in the spinal cord and have implications for the interpretation of the labeling obtained when using WGA-HRP.

Fine structure and synaptic connections of the spinal dorsal root terminals in the turtle Chrysemys d'orbigny

The synaptology of dorsal root afferents in the spinal cord of the turtle Chrysemys d'orbigny was analyzed employing light and serial section reconstruction electron microscope (EM) techniques. Horseradish peroxidase-marked dorsal root terminals were found distributed in four ipsilateral dorso-ventral synaptic fields termed here Ia, Ib, II and III. Commisural dorsal root fibers were found projecting to the contralateral dorsal horn. When observed under the EM the labeled dorsal root endings showed mitochondria and cumuli of large vesicles. Complex synaptic arrays consisting of dorsal root endings and dendritic processes occur in the peripheral synaptic fields (Ia, Ib and II); more simple axo-dentritic junctions predominate at more ventral levels. The EM images suggest that the sensory terminals not only behave as pre-synaptic elements, but also as post-synaptic ones with respect to the dorsal horn neurons. In addition, the occurrence of numerous axo-axonic symmetrical junctions enable us to speculate about direct interactions between primary afferents.

Further evidence in support of taurine as a mediator of synaptic transmission in the frog spinal cord

It has been reported that 6-aminomethyl-3-methyl-4H,1,2,4-benzothiadiazine-1,1-dioxide (AMBD, TAG) is a specific blocker of taurine and β-alanine responses in the central nervous system. We have re-examined the effect of AMBD on amino acid and synaptically evoked responses recorded from isolated hemisected frog spinal cords by means of the sucrose gap technique. When indirect responses were blocked by adding tetrodotoxin (0.2 μM) or manganese chloride (2 mM) to the normal Ringer solution, AMBD (0.01–0.5 mM) selectively antagonized taurine, β-alanine, hypotaurine and kojic amine evoked depolarizations of primary afferents at their intramedullary part (dorsal root terminals, DRT) and on dorsal root ganglia (DRG), without significantly affecting responses to glutamate (on DRT), glycine (on DRT) or GABA (on DRT and DRG). Depolarizing responses to taurine and β-alanine (1 mM) were depressed by up to 50% with 0.1 mM AMBD and often completely antagonized with 0.25 mM AMBD. In normal Ringer solution, AMBD selectively antagonized the dorsal root potential evoked by ventral root stimulation (VR-DRP, threshold at 0.02 mM AMBD, 90% block with 0.25 mM); other synaptic potentials increased in duration and/or amplitude, demonstrating a strong convulsant effect of AMBD. Thus, the depolarizing responses of taurine, β-lanine and hypotaurine on primary afferents are pharmacologically indistinguishable from the VR-DRP. These results are in agreement with the proposal that taurine or a taurine-like substance (possibly β-alanine or hypotaurine) is the mediator of VR-DRP in amphibian spinal cord.

Grafts of Pacinian corpuscles reinnervated by dorsal root axons

In adult rats, a piece of the crural interosseous nerve with several Pacinian corpuscles attached was removed from the crural region, autotransplanted onto the surface of the lumbar spinal cord and connected with the peripheral stump of a transected dorsal root. From 10 days up to 6 months after the operation, the grafts were investigated by light and electron microscopy. The regenerating dorsal root axons grew along the grafted nerves into the attached Pacinian corpuscles. By 1–2 months after the operation, the nerves and their branches became almost completely reinnervated by myelinated and unmyelinated dorsal root axons. In a sample of corpuscles examined 2–6 months after grafting, 75% of corpuscles were found reinnervated; each of them was supplied by 1–5 large myelinated axons that formed multiple axon terminals in the inner core. The maximal number of axonal profiles found in a transverse section through different levels of the inner core varied, in individual corpuscles, from 3 to 17 axons and terminals. The dorsal root terminals formed in the grafted corpuscles were mainly filled with mitochondria and resembled peripheral sensory endings. In some instances, the newly formed endings developed lateral processes and membrane specializations characteristi for peripheral Pacinian terminals. Thus regenerating dorsal root axons recognize a grafted peripheral mechanoreceptor as their target and reinnervate it with axon terminals, most of them structurally transformed into peripheral sensory endings.

Replacement of synaptic terminals in lamina II and Clarke's nucleus after unilateral lumbosacral dorsal rhizotomy in adult cats

Evidence from previous light-microscopic studies suggested that lumbosacral dorsal rhizotomy in cats elicits sprouting of converging undamaged systems into partially deafferented Clarke's nucleus and lamina II. We therefore applied quantitative electron-microscopic methods to determine whether this sprouting is associated with replacement of synaptic terminals (reactive reinnervation). We used stereological and morphometric methods to estimate terminal number per cross section in right and left lamina II and Clarke's nucleus in adult cats after acute and chronic unilateral (right-sided) lumbosacral deafferentation. Planimetric measurements of area indicated no significant shrinkage of either region as a result of the deafferentation; an increase in area occupied by glial cytoplasm (gliosis) equaled the decrease occupied by axonal components. The gliosis appears to persist indefinitely, although the degenerative debris stainable with conventional light-microscopic methods does not persist. Analysis of the synaptic population of lamina II reveals that the large central or "scalloped" terminals comprise a substantial fraction (greater than 40%) of the total area occupied by terminals in control material and that this population is largely lost upon deafferentation, leaving a large population of small terminals with spherical vesicles. Nevertheless, estimates of total terminal number indicate no difference between control and deafferented lamina II, suggesting a rapid and virtually complete replacement of lost dorsal root terminals by small terminals containing spherical vesicles. Terminal number in Clarke's nucleus also remains constant despite the loss of the dorsal root input. We conclude that there is also a virtually complete and rapid replacement of lost terminals in Clarke's nucleus by terminals containing spherical vesicles. These data provide an example of a case in which axonal sprouting demonstrated with light-microscopic methods is associated with electron-microscopic evidence of reactive reinnervation.

Dorsal root afferents contact migrating motoneurons in the developing frog spinal cord

Dorsal and ventral roots of Rana catesbeiana tadpole lumbar spinal cord were labeled with horseradish peroxidase during development of the hindlimb. Labeled motoneurons in the process of migrating were found in the region of the developing terminal arbors of dorsal root axons. Electron microscopy showed that some of these dorsal root terminals contacted the migrating motoneurons.

Catecholamine effects on frog dorsal root terminals

Dopamine, norepinephrine and epinephrine applied to the isolated superfused frog spinal cord had complex effects on the terminals of primary afferent fibers. The most cosistent finding was a slow hyperpolarization of terminals with lower concentrations (10 μM or lower), but depolarization either following or admixed with the hyperpolarization were seen. These were particularly prominent when the catecholamines were applied in high concentrations or for prolonged periods of time. A part of the response of afferent terminals appears to be indirect since the potential changes were reduced following exposure of the cord to tetrodotoxin, Mn 2+, or mephenesin. The hyperpolarizations were augmented by imipramine, a known inhibitor of catecholamine uptake. These observations are consistent with a role of catecholamines in the processing of sensory input in the spinal cord.

Periterminal synaptology of dorsal root glomerular terminals in the substantia gelatinosa of the spinal cord in the rhesus monkey.

The synaptic glomerular complexes surrounding dorsal root terminals in the substantia gelatinosa were reconstructed from six sets of 50--140 gapless ultrathin serial sections prepared in the transverse plane of the spinal cord in adult rhesus monkeys. All three types of glomerular terminals described in the preceding paper (Knyihar-Csillik et al., '82) were identified: (1) DSA (endings of superficial collaterals); (2) LDCV (endings of marginal collaterals); and (3) RSV (endings of deep collaterals). Each type of terminal forms a glomerular complex which invariably includes presynaptic dendrites which are intercalated between primary terminals and the postsynaptic (conventional) dendrite. Since the latter also receives direct input from the primary sensory terminal the synaptic organization assumes triadic arrangements, suggesting that primary afferent impulses may be subjected to a postsynaptic modulation through inhibitory action of presynaptic dendrites. In glomeruli with DSA as the central element, several triadic systems are usually interrelated, possibly as a structural basis for prolonged retardation of impulses. Adjacent glomeruli, containing DSA and LDCV terminals, are coupled together by a series of triadic systems fed by DSA terminals enabling association between superficial and marginal collaterals. RSV terminals are presynaptic to somata and dendrites of substantia gelatinosa cells that presumably exert inhibition upon terminals of all three kinds of primary sensory collaterals. In addition RSV terminals are postsynaptic to numerous F boutons which presumably derive mainly from axons of substantia gelatinosa cells; similar F boutons impinge upon presynaptic and other dendrites surrounding DSA terminals. The complicated but orderly synaptic architecture of these types of primary afferents may be regarded as a structural basis for first-order analysis and modulation of the nociceptive information within the primate central nervous system.

Hyperpolarization of frog primary afferent fibres caused by activation of a sodium pump.

1. In the isolated frog spinal cord repetitive stimulation of a lumbar dorsal root produced a sustained negative potential recorded from an adjacent inactive dorsal root by sucrose gap techniques. This negative potential was followed by a positive potential, an indication that the dorsal root terminals were hyperpolarized. Increasing the duration of the tetanus applied to the active root increased the amplitude and duration of the after-hyperpolarization which could be up to 6 mV and 3 min respectively. 2. The hyperpolarization presumably reflected an increased rate of active sodium pumping. Since it was reversibly reduced by metabolic inhibitors (dinitrophenol, NaCN) and cooling (Q10, 2 . 6) it was clearly dependent upon intact metabolic activity. In addition, a variety of procedures used to inhibit sodium pumps (including application of ouabain, elimination of potassium from the superfusate, and partial substitution of lithium for sodium ions) significantly and reversibly decreased the potential. 3. The hyperpolarization was not dependent upon intact chemical synaptic transmission since it could survive prolonged immersion of the cord in Ringer solution containing manganese or magnesium ions. 4. It is suggested that the hyperpolarization of inactive fibres resulted from a decreased extracellular potassium concentration in the dorsal horn produced as a result of a pumping mechanism which extruded sodium and transported potassium inwards by dorsal root fibres directly activated by the tetanus.

Motilin excites neurons in the cerebral cortex and spinal cord

The 22 amino acid polypeptide motilin was tested by iontophoretic application onto neurons in the rat cerebral cortex and by perfusion over the isolated hemisected toad spinal cord. Motilin (25–150 nA) excited identified cortico-spinal neurons and other deep spontaneously firing cortical icells. Excitation developed relatively rapidly and lasted for up to 60 sec after the application. Motilin was a potent excitant (threshold concentration (2.5 × 10 −9 M) of neurons in the amphibian spinal cord, eliciting a depolarization of dorsal root terminals and motoneurons. Its effects were substantially reduced after tetrodotoxin, suggesting a primary site of action on spinal cord interneurons.

Actions of various gastrointestinal peptides on the isolated amphibian spinal cord.

The effects of a number of peptides which are found in the gastrointestinal tract have been ascertained on the direct current recorded dorsal and ventral root responses of the isolated hemisected toad spinal cord. Motilin, substance P, bombesin, neurotensin, and thyrotropin releasing hormone had potent depolarizing actions on dorsal root terminals and motoneurons. These substances evoked discernable effects at concentrations as low as 10--7 M, or even lower with motilin. The effects of motilin, neurotensin, and thyrotropin-releasing hormone were greatly reduced or abolished by perfusion of the preparation with tetrodotoxin. Adrenocorticotrophic hormone, secretin, and pancreozymin (cholecystokinin) also depolarized dorsal root terminals and motoneurons. The effects of secretin and cholecystokinin were not abolished by tetrodotoxin. Leu- and Met-enkephalin had weak hyperpolarizing actions on the dorsal and ventral root potentials of repetitively stimulated preparations. Gastrin, gastric inhibitory peptide, glucagon, and somatostatin had no apparent effects on the responses of the preparation. Angiotensin and vasopressin both had rather weak depolarizing effects on the dorsal and ventral roots.

Action of biogenic amines on the isolated toad spinal cord

1. The effects of noradrenaline, adrenaline, dopamine and 5-hydroxytryptamine have been tested on isolated hemisected spinal cords in which synaptic transmission or action potential propagation were blocked by perfusion with high Mg 2+—or tetrodotoxin—containing solutions respectively. 2. The catecholamines at a concentration of 10 −3M consistently depolarized dorsal root terminals, and in most preparations elicited a small depolarization of motoneurons. At lower concentrations their effects were less consistent. The threshold concentration for discernable effects on the dorsal roots was ∼10 −4M. 3. Desensitization to its own effects and those of the other catecholamines occurred following catecholamine application. Recovery from desensitization took up to 60 min. 4. Noradrenaline evoked depolarizations of both roots were antagonized by phentolamine (2 × 10 −5M) and propranolol (10 −4M). There may be both α- and β-receptors on amphibian neurons and axon terminals. 5. 5-Hydroxytryptamine (10 −4M) consistently depolarized dorsal root terminals, usually with a small depolarization of the ventral roots. Higher concentrations elicited pronounced depolarizations in both roots. Desensitization to the effects of 5-hydroxytryptamine was observed with repeated applications. Cross-desensitization between the indole-amine and catecholamines was not observed.

Vasoactive intestinal polypeptide excitation of central neurons

Vasoactive intestinal polypeptide (VIP) was tested on neurons in the rat sensory motor cerebral cortex and on the isolated hemisected toad spinal cord. Iontophoretically applied VIP excited deep, spontaneously active cortical neurons, including identified corticospinal neurons. The excitation had a latency of onset varying from several seconds to over 1 min and often lasted for a minute or longer after cessation of the application. Desensitization of the effect occurred with repeated applications. VIP caused a depolarization of motoneurons and dorsal root terminals in the isolated amphibian spinal cord. Threshold for this effect was about 10(-6) M. The effects of VIP on both preparations were comparable with those of another peptide, substance P.

The actions of adenosine and various nucleosides and nucleotides on the isolated toad spinal cord

1. 1. The effects of nucleosides and nucleotides of several purines and pyrimidines have been ascertained on the a.c. and d.c. recorded dorsal and ventral root responses of isolated, hemisected, perfused toad spinal cords. 2. 2. On preparations subjected to repetitive dorsal root stimulation, adenosine and adenosine 5′-monophosphate, 5′-diphosphate and 5′-triphosphate (2 × 10 −4−10 −2 M) hyperpolarized both dorsal and ventral roots. Dorsal root-evoked dorsal and ventral root potentials were reduced in duration but not amplitude. These effects were antagonized by theophylline and caffeine. 3. 3. Adenosine 5′-triphosphate and, to a lesser extent, adenosine 5′-diphosphate, at concentrations of 10 −3 M or higher frequently depolarized one or both roots. This effect was potentiated by caffeine and theophylline. 4. 4. Guanosine 5′-monophosphate, 5′-diphosphate and 5′-triphosphate (10 −3−10 −2 M) hyperpolarized both roots and reduced both the amplitude and duration of dorsal and ventral root potentials evoked by dorsal root stimulation. These effects were only weakly antagonized by caffeine and theophylline. Depolarizations of the roots were sometimes observed with guanosine and its 5′-di- and triphosphates. 5. 5. Adenosine 3′,5′-cyclic monophosphate and guanosine 3′,5′-cyclic monophosphate had no effect at concentrations of up to 10 −2 M on the responses and polarization levels of the dorsal and ventral roots. 6. 6. Inosine 5′-monophosphate at high concentrations (10 −3−10 −2 M) had a weak hyperpolarizing action on the ventral root potential. Cytidine, thymidine, uridine and xanthosine and their 5′-monophosphates had no effect at concentrations of up to 10 −2 M. The 5′-triphosphates of these substances all depolarized dorsal and ventral roots. 7. 7. After perfusion with magnesium-containing solutions, adenine nucleotides and uridine 5′-triphosphate depolarized the dorsal roots but had no consistent effect on ventral root polarization levels. 8. 8. The sodium and dicalcium salts of adenosine 5′-triphosphate had equipotent depolarizing actions. The excitant actions of the nucleotides are therefore unlikely to be due to calcium chelation. 9. 9. In conclusion, it appears that adenosine and the adenine nucleotides can have depressant and hyperpolarizing actions on dorsal root terminals and motoneurons of repetitively stimulated preparations. It is suggested that these effects result from a reduction in the release of excitatory transmitter. The 5′-polyphosphates of all the purines and pyrimidines tested can have an excitatory action, the nature of which is currently uncertain.

Electron microscopic identification of postsynaptic dorsal root terminals: a possible substrate of dorsal root potentials in the frog spinal cord

Dorsal root fibers were labeled with cobaltous chloride iontophoresis for electron microscopic investigations. In the base of the dorsal horn, where most of the coarser collaterals of dorsal root fibers terminate, many dorsal root terminals were found in postsynaptic relation to synapsing profiles. According to their morphological characteristics, three kinds of presynaptic terminals could be discerned in these complex synapses: axon terminals with spheric vesicles, axon terminals with flattered vesicles and presynaptic dendrites. These latter terminals contained relatively few flattened vesicles accumulated adjacent to a short synaptic articulation surface, and they were rich in cytoplasmic organelles. The functional significance of these structural specializations in the mediation of dorsal root potentials and recurrent inhibition is discussed.

The origin of spinocerebellar pathways. I. The nucleus cervicalis centralis of the cranial cervical spinal cord

AbstractThe origin of spinocerebellar projections from the cranial cervi cal spinal cord was studied in neonatal dogs following cerebellar ablations or the injection of horseradish peroxidase (HRP) into the cerebellum. Cerebellar ablations produced distinct retrograde changes of three types: type I response or cen tral chromatolysis; type II response or ghost cells; and type 111 response or neu ronal loss and gliosis. These ablation‐induced changes were evident in the cells of the nucleus cervicalis centralis. The results of hemicerebellectomy, unilateral cer ebellar pedunculotomy, and cordotomy established that the cells of this nucleus project contralaterally to the cerebellum. Cytoarchitectonic study revealed that this nucleus is composed of clustered medium‐sized, multipolar, chromatophilic neurons located in the zona intermedia of the C1‐C4 segments. The retrograde la beling of central cervical neurons following cerebellar injections of HRP con firmed the results obtained with the axonal reaction.Anterograde fiber degeneration was present in the central cervical nucleus following dorsal rhizotomy in the dog and cat. These experiments demonstrated dorsal root fibers among the somata of central cervical neurons rendered chromatolytic following cerebellectomy in the dog. Electron microscopic evidence is presented for the presence of degenerated dorsal root terminals upon the dendrites and somata of central cervical neurons.The nucleus cervicalis centralis forms a direct link between neck afferents and the cerebellum by way of a crossed cervicospinocerebellar pathway. Evidence could not be found for the presence of a spinocerebellar projection originating in the nucleus ceruicalis lateralis.

Further studies on glutamate antagonists in the central nervous system

1. 1. Three diaminomonocarboxylic acids which reportedly antagonize the increase in cyclic 3′,5′-adenosine monophosphate in brain slices induced by l-glutamate were tested as glutamate antagonists on the toad ( Bufo marinus) spinal cord and on cerebral cortical neurones in the rat. 2. 2. l-2,4-diaminobutyric acid, dl-2,3-diaminopropionic acid and l-ornithine depressed dorsal root-evoked responses recorded from the dorsal and ventral roots of the amphibian spinal cord and depolarized dorsal root terminals. Associated with these effects there was a reduction in glutamate-evoked depolarization of dorsal root terminals and spinal motoneurones. 3. 3. The three diaminocarboxylic acids depressed the spontaneous, and the l-glutamate-, l-aspartate-and acetylcholine-evoked firing of cerebral cortical neurones. 4. 4. It is concluded that the diaminocarboxylic acids exhibit a similar, albeit less potent, type of action to the monoaminocarboxylic acids, γ-aminobutyric acid (GABA) and β-alanine. They are unlikely to be useful as glutamate antagonists in neuropharmacological experiments.