Dorsal spinocerebellar tract neuronal activity in the intact chronic cat

The ability to electrophysiologically identify the axonal projections of lumbar neurons recorded in chronic unanesthetized intact awake animals is a formidable but essential requirement toward understanding ascending sensory transmission under naturally occurring conditions. Chronic immobilization procedures previously introduced by Morales et al. (1981) for intracellular studies of motoneurons are modified and then integrated with procedures for antidromic cellular identification and extracellular recording of upper (or lower) dorsal lumbar spinocerebellar tract (DSCT) neuronal activity, in conjunction with behavioral state recording and drug microiontophoresis. These implant procedures provide up to 6 months of stable recording conditions and, when combined with other techniques, allow individual DSCT neurons to be monitored over multiple cycles of sleep and wakefulness, following the induction into and recovery from barbiturate anesthesia and/or during the juxtacellular microiontophoretic ejection of inhibitory or excitatory amino acid neurotransmitters. The combination of such techniques allows a comprehensive examination of synaptic transmission through the DSCT and other lumbar sensory pathways in the intact normally respiring cat and its modulation during the general anesthetic state. These techniques permit investigations of the supraspinal controls impinging on lumbar sensory tract neurons during wakefulness and other behavioral states such as active sleep.     

Secondary vestibulo-oculomotor projections in larval sea lamprey: anterior octavomotor nucleus

The ventral octavolateral area of lampreys contains three nuclei: the anterior, intermediate and posterior octavomotor nuclei, formed of large neurons that are contacted by thick primary vestibular fibres. We used horseradish peroxidase (HRP) or fluorescein-dextran-amine (FDA) labelling to study the projections of the anterior octavomotor nucleus (AON) in the larval sea lamprey, Petromyzon marinus. The tracers were injected either in the AON, the oculomotor nucleus or the rostralmost spinal cord. HRP injection in the AON labelled thick axons that coursed to the basal mesencephalic tegmentum, where most decussate and project to the oculomotor nucleus and the third Müller cell. Electron microscopy confirmed that AON axons contact with the contralateral third Müller cell and with oculomotor neurons. Some AON axons run in the mesencephalic tegmentum and the ventral diencephalon. An AON axon was observed to run close to the axon of the contralateral third Müller cell, establishing what appeared to be en passant contacts. HRP injection in the AON also revealed commissural fibres projecting to the contralateral octavolateral area. HRP or FDA injections in the oculomotor nucleus labelled both large and small neurons of the AON, mostly contralateral to the injection site, as well as of cells in the intermediate octavomotor nucleus, mainly ipsilateral. HRP injection in the AON or in the rostral spinal cord did not reveal any projections from the AON to the spinal cord. Our results indicate that the pattern of octavo-oculomotor connections in the lamprey is different from that observed in other vertebrates.     

Effects of neurotransplants and BDNF on the survival and regeneration of injured adult spinal motoneurons

We compared the effects of peripheral nerve grafts, embryonic spinal cord transplants and brain-derived neurotrophic factor (BDNF) on the survival and axon regeneration of adult rat spinal motor neurons undergoing retrograde degeneration after ventral root avulsion. Following implantation into the dorsolateral funiculus of the injured spinal cord segment, neither a peripheral nerve graft nor a combination of peripheral nerve graft with embryonic spinal cord transplant could prevent the retrograde motor neuron degeneration induced by ventral root avulsion. However, intrathecal infusion of BDNF promoted long-term survival of the lesioned motor neurons and induced abundant motor axon regeneration from the avulsion zone along the spinal cord surface towards the BDNF source. A combination of ventral root reconstitution and BDNF treatment might therefore be a promising means for the support of both motor neuron survival and guided motor axon regeneration after ventral root lesions.     

Cirrhosis and muscle cramps: evidence of a causal relationship

Enkephalin-like immunoreactivity (ENK-LI) was found throughout the spinal cord of the long-tailed ray Himantura fai. The densest ENK-LI was in the superficial portion of lamina A of the dorsal horn. Lamina B and the deeper parts of lamina A contained radially oriented, labelled fibres. Laminae C, D, and E contained many longitudinally oriented fascicles which were surrounded by a reticulum of transversely oriented, labelled fibres, some of which projected into the ventral and lateral funiculi. Labelled fibres were found in the dorsal commissure and around the central canal, but the later did not cross the midline. One-third of all enkephalinergic cells were found throughout laminae A and B, while two-thirds were located in the medial half of C, D, and E. Occasionally a labelled cell was located in the lateral funiculus. The ventral horn (laminae F and G) contained many enkephalinergic fibres but no labelled nuclei. A few dorsal column axons contained ENK-LI. In the lateral funiculus there were two groups of labelled axons, a superficial, dorsolateral group, and a deeper group, occupying a crescent-shaped region. The ventral funiculus also contained many labelled axons. The central projection of the dorsal root passed through the substantia gelatinosa and divided into rostrally and caudally projecting fascicles within lamina C. The root, and these fascicles, both lacked ENK-LI. In contrast, the fascicles in laminae D and E did contain enkephalinergic fibres. The origin of the various fibre systems and the role of enkephalin in the regulation of sensory processing and motor output are discussed.     

Red nucleus stimulation inhibits within the inferior olive

Red nucleus stimulation inhibits within the inferior olive. J. Neurophysiol. 80: 3127-3136, 1998. In the anesthetized cat, electrical stimulation of the magnocellular red nucleus (RNm) inhibits responses of rostral dorsal accessory olive (rDAO) neurons to cutaneous stimulation. We tested the hypothesis that RNm-mediated inhibition occurs within the inferior olive by using stimulation of the ventral funiculus (VF) of the spinal cord in place of cutaneous stimulation of the hindlimb. Fibers in the VF terminate on hindlimb rDAO neurons, so inhibition of this input would have to occur within the olive. rDAO responses elicited by VF stimulation were inhibited by prior stimulation of the RNm, indicating that inhibition occurs within the olive. In contrast, evoked potentials recorded from the VF or dorsal columns following hindlimb stimulation were not affected by prior stimulation of RNm, indicating that stimulation of the RNm does not inhibit olivary afferents at spinal levels. RNm stimulation that inhibited rDAO responses had little effect on evoked somatosensory responses in thalamus, indicating that inhibition generated by activity in RNm may be specific to rDAO. To test limb specificity of RNm-mediated inhibition, conditioning stimulation was applied to the dorsolateral funiculus at thoracic levels, which selectively activates RNm neurons projecting to the lumbar cord. Stimulation at thoracic levels inhibited evoked responses from hindlimb but not forelimb regions of rDAO, suggesting that inhibitory effects of RNm activity are limb specific. Several studies have reported that olivary neurons have reduced sensitivity to peripheral stimulation during movement; it is likely that RNm-mediated inhibition occurring within the olive contributes to this reduction of sensitivity. Inhibition of rDAO responses by descending motor pathways appears to be a salient feature of olivary function.     

Tracing the auditory pathways to electrophysiologically characterized neurons with HRP and Fos double-labeling technique

By combining HRP histochemistry with Fos immunocytochemistry, we demonstrate in this study that electrophysiologically characterized auditory neurons can be double-labeled with HRP and Fos after iontophoretic injection of HRP into the recording site. Neurons which projected fibers to the recording site were labeled with HRP and were Fos-like immunoreactive. This double-labeling technique in combination with electrophysiological recording offers the possibility to determine the fiber projections between sound-activated neurons which are identified either electrophysiologically and/or immunocytochemically.     

Differential effects of morphine on corneal-responsive neurons in rostral versus caudal regions of spinal trigeminal nucleus in the rat

The initial processing of corneal sensory input in the rat occurs in two distinct regions of the spinal trigeminal nucleus, at the subnucleus interpolaris/caudalis transition (Vi/Vc) and in laminae I-II at the subnucleus caudalis/spinal cord transition (Vc/C1). Extracellular recording was used to compare the effects of morphine on the evoked activity of corneal-responsive neurons located in these two regions. Neurons also were characterized by cutaneous receptive field properties and parabrachial area (PBA) projection status. Electrical corneal stimulation-evoked activity of most (10/13) neurons at the Vi/Vc transition region was increased [146 +/- 16% (mean +/- SE) of control, P < 0.025] after systemic morphine and reduced after naloxone. None of the Vi/Vc corneal units were inhibited by morphine. By contrast, all corneal neurons recorded at the Vc/C1 transition region displayed a naloxone-reversible decrease (55 +/- 10% of control, P < 0.001) in evoked activity after morphine. None of 13 Vi/Vc corneal units and 7 of 8 Vc/C1 corneal units tested projected to the PBA. To determine if the Vc/C1 transition acted as a relay for the effect of intravenous morphine on corneal stimulation-evoked activity of Vi/Vc units, morphine was applied topically to the dorsal brain stem surface overlying the Vc/C1 transition. Local microinjection of morphine at the Vc/C1 transition increased the evoked activity of 4 Vi/Vc neurons, inhibited that of 2 neurons, and did not affect the remaining 12 corneal neurons tested. In conclusion, the distinctive effects of morphine on Vi/Vc and Vc/C1 neurons support the hypothesis that these two neuronal groups contribute to different aspects of corneal sensory processing such as pain sensation, autonomic reflex responses, and recruitment of descending controls.     

Intracortical arborizations and receptive fields of identified ventrobasal thalamocortical afferents to the primary somatic sensory cortex in the cat

The intracortical arborizations of neurons from the ventroposterolateral thalamic nucleus (VPL) in the cat were studied by intraaxonal injections of horseradish peroxidase (HRP) following identification of their receptive fields. In the primary somatic sensory cortex (SI) VPL cells terminated in different cytoarchitectonic areas according to their receptive field modality. Fibers excited by deep tissue or joint rotation arborized preferentially in area 3a. Those responding tonically to cutaneous stimuli were located in the anterior part of area 3b; hairdriven cells terminated in area 3b and in the rostral pole of area 1. All fibers had a similar laminar distribution within SI. Axons terminated mostly in layers VI, iV, and the lower part of layer III. None terminated in layers I and II. Most terminal arbors were oriented along the mediolateral axis of the brain. The main arborization of a single VPL cell formed a bush of about 500 micrometers in diameter. some fibers generated two such bushes with an uninvaded region of about 300 micrometer between them. It is proposed that this patchy organization underlies in part the columnar organization of areas SI. Many VPL cells had secondary projection sites in SI. These were issued from smaller-sized collaterals and were located in a different cytoarchitectonic area than that of the main terminal plexuses. A significant number of these collaterals projected to area 4, Insufficient filling of the collaterals by HRP prevented a more complete characterization of the secondary arbors.     

Effect of nitrous oxide on spinal dorsal horn WDR neuronal activity in cats

The effects of nitrous oxide (75%) on the spinal dorsal born wide dynamic range (WDR) neuronal activity were studied in either spinal cord intact or spinal cord-transected cats. Extracellular activity was recorded in the dorsal horn from single WDR neurons responding to noxious and non-noxious stimuli applied to the cutaneous receptive fields on the left bind foot pads of intact or decerebrate, spinal cord-transected (L 1-2) cats. The experiment was divided into four sections as follows: (1) When 10 micrograms of bradykinin (BK) was injected into the femoral artery ipsilateral to the recording site as the noxious test stimulus in the spinal cord-transected cat, all of 6 WDR neurons gave excitatory responses which were not depressed by 75% nitrous oxide. (2) When the injection of 10 micrograms of BK into the femoral artery ipsilateral to the recording site was used in the spinal cord-intact cat, 6 of 15 WDR neurons (40%) gave excitatory responses, which were significantly depressed by 75% nitrous oxide, and 9 of 15 WDR neurons (60%) gave inhibitory responses, which were not affected by 75% nitrous oxide. (3) When 10 micrograms of bradykinin (BK) was injected into the femoral artery contralateral to the recording site as the noxious test stimulus in the spinal cord transected cat, 6 of 12 WDR neurons gave excitatory reasons, which were not depressed by 75% nitrous oxide. (4) When the injection of 10 micrograms of BK into the femoral artery contralateral to the recording site was used in the spinal cord-intact cat, 6 of 6 WDR neurons (100%) gave responses, which were affected by 75% nitrous oxide. We have observed that nitrous oxide reduces the excitation and inhibition of dorsal born WDR neuronal activities induced by BK injection in spinal cord-intact cats, but does not reduce the excitation of those in spinal cord-transected cats. This finding confirmed that the antinociceptive effect of nitrous oxide might be modulated by supraspinal descending inhibitory control systems. In addition our result showed that the supraspinal effect of nitrous oxide was mediated not only by an increase but also a decrease in a supraspinal descending inhibition.     

Dorsal horn projection targets of ON and OFF cells in the rostral ventromedial medulla

1. The rostral ventromedial medulla (RVM) participates in the modulation of nociceptive transmission by spinal cord neurons. Previous anatomic studies have demonstrated that RVM neurons project to laminae I, II, and V of the dorsal horn; laminae VII and VIII of the intermediate and ventral horns; the intermediolateral column; and lamina X. The RVM contains at least three physiologically defined classes of neurons, two of which, the ON and the OFF cells, have been implicated in nociceptive modulation. Because these cells classes are intermingled in the RVM, it has not been possible to determine the spinal laminar projection targets of ON and OFF cells by anatomic methods. Therefore in the current study we employed antidromic microstimulation methods to determine the laminar projections of two of the three classes of RVM neurons, the ON and the OFF cells. 2. In lightly anesthetized (with methohexital sodium) rats, single-unit extracellular recordings were made from 48 RVM neurons that were physiologically characterized as ON (30) or OFF (18) cells. The recording locations of 45 of these neurons were recovered. Thirty-seven were found in the nucleus raphe magnus and eight were located near its dorsal and lateral borders. 3. Thirty-two physiologically identified RVM neurons (18 ON and 14 OFF cells) were antidromically activated from the cervical spinal cord using a monopolar stimulating electrode. The stimulating electrode was moved systematically in the white matter until antidromic activation could be produced with currents of < or = 20 microA (6.1 +/- 0.7 microA, mean +/- SE). The points from which minimum currents were required to antidromically activate the neurons were located mainly in the ipsilateral dorsolateral funiculus (DLF) (27 of 32). In a few cases, lowest antidromic threshold currents were found near the border between the DLF and ventrolateral funiculus (VLF) or, rarely, in the VLF itself. In these cases, the cell recordings were found to be near the dorsal boundary of the RVM. 4. While one electrode was used to stimulate the parent axon in the lateral funiculus, a second was used to explore the gray matter for the presence of collateral branches. The identification of a branch was initially determined by an increase in antidromic latency. At the same rostrocaudal plane of the spinal cord, stimulation of the DLF induced an antidromic spike that invaded the neuron earlier than the antidromic spike elicited by stimulation in the gray matter. Collateral branches were confirmed by establishing that the location of the minimum threshold point for antidromic activation of the neurons from the second electrode was in the gray matter, that the minimum current required to antidromically activate the neuron from that point was too low to activate the parent axon in the DLF, and that a collision occurred between the spikes induced by the two stimulating electrodes. 5. In 17 cases, physiologically identified RVM neurons (10 ON and 7 OFF cells) were antidromically activated from the gray matter of the cervical spinal cord using a current of 8.4 +/- 2.1 (SE) microA. Minimum threshold points for antidromic activation were found in laminae I-II (3 ON and 4 OFF cells), lamina V (5 ON and 6 OFF cells), and regions ventral to the lateral reticulated area (3 ON and 2 OFF cells) of the gray matter. As indicated by these numbers, some neurons were antidromically activated from more than one gray matter region. In general, all OFF cells and 9 of 10 ON cells were antidromically activated from low threshold points in either laminae I-II or lamina V. 6. In six cases, neurons were activated from separate points located in two or three different laminae of the gray matter. Three OFF cells were activated from laminae I-II and V, one OFF cell and one ON cell were activated from lamina V and from more ventral points, and one ON cell was activated from laminae I-II and from points ventral to lamina V.     

Post-traumatic reconnection of the cervical spinal cord with skeletal striated muscles. Study in adult rats and marmosets

In an attempt at repairing the injured spinal cord of adult mammals (rat, dog and marmoset) and its damaged muscular connections, we are currently using: 1) peripheral nerve autografts (PNG), containing Schwann cells, to trigger and direct axonal regrowth from host and/or transplanted motoneurons towards denervated muscular targets; 2) foetal spinal cord transplants to replace lost neurons. In adult rats and marmosets, a PNG bridge was used to joint the injured cervical spinal cord to a denervated skeletal muscle (longissimus atlantis [rat] or biceps brachii [rat and marmoset]). The spinal lesion was obtained by the implantation procedure of the PNG. After a post-operative delay ranging from 2 to 22 months, the animals were checked electrophysiologically for functional muscular reconnection and processed for a morphological study including retrograde axonal tracing (HRP, Fast Blue, True Blue), histochemistry (AChE, ATPase), immunocytochemistry (ChAT) and EM. It was thus demonstrated that host motoneurons of the cervical enlargement could extend axons all the way through the PNG bridge as: a) in anaesthetized animals, contraction of the reconnected muscle could be obtained by electrical stimulation of the grafted nerve; b) the retrograde axonal tracing studies indicated that a great number of host cervical neurons extended axons into the PNG bridge up to the muscle; c) many of them were assumed to be motoneurons (double labelling with True Blue and an antibody against ChAT); and even alpha-motoneurons (type C axosomatic synapses in HRP labelled neurons seen in EM in the rat); d) numerous ectopic endplates were seen around the intramuscular tip of the PNG. In larger (cavitation) spinal lesions (rat), foetal motoneurons contained in E14 spinal cord transplants could similarly grow axons through PNG bridges up to the reconnected muscle. Taking all these data into account, it can be concluded that neural transplants are interesting tools for evaluating both the plasticity and the repair capacities of the mammalian spinal cord and of its muscular connections.     

Systolic intervals in disorders of intraventricular conduction]

After HRP injections into the octopus cell area of the cat cochlear nucleus, only periolivary neurons of the superior olivary complex (SOC) reacted. Elongate neurons in the lateral periolivary nuclei (ipsilateral to the injection) and multipolar neurons in ventromedial periolivary regions (contralateral to the injection) contained granules. No neurons in the main SOC nuclei or higher auditory nuclei reacted, despite a wide range of HRP concentrations. Thus, neurons from the SOC to the octopus cell area of the cochlear nucleus seem to be entirely periolivary and not entirely equivalent to neurons providing collaterals to the olivocochlear bundle.     

Anatomical study of spinobulbar neurons in lampreys

The present study was aimed at identifying spinal neurons ascending to the brainstem outside the dorsal columns in the lamprey. Two retrograde tracers (cobalt-lysine and horseradish peroxidase [HRP]) were injected in the brainstem or rostral spinal cord in vivo or in vitro. Labeled cells were distributed bilaterally with a contralateral dominance, along the whole rostrocaudal extent of the spinal cord. The density of cells markedly decreased rostrocaudally. Several classes of brainstem-projecting neurons were identified. Most cells with a short axon were small and formed columns, in the dorsolateral and ventrolateral gray matter, at the transition between the rhombencephalon and the spinal cord. Dorsal elongated cells were spindle shaped, located medially, in the first two spinal segments. Lateral elongated cells were medium to large size neurons, located in the intermediate and lateral gray matter, mainly contralateral to the injection site. Their axon emerging from the lateral part of the soma crossed the midline, ventral to the central canal. These cells were present throughout the rostral spinal cord. Cells were also labeled in the lateral white matter. Some of them had the typical dendritic arborizations of edge cells (intraspinal stretch receptor neurons) and were located in the most rostral segments, bilaterally. Other medium to large size neurons were identified dorsal and medial to most of the edge cells. We suggest that at least the group of lateral elongated cells exhibits rhythmic membrane potential oscillations during fictive locomotion. These cells may, together with the rostral edge cells, be responsible for the locomotor-related modulation of activity in reticulospinal and vestibulospinal neurons.     

Distribution and connections of inspiratory premotor neurons in the brainstem of the pigeon (Columba livia)

We have recorded extracellular, inspiratory-related (IR) unit activity in the medulla at locations corresponding to those of neurons retrogradely labeled by injections of retrograde tracers in the lower brachial and upper thoracic spinal cord, injections that covered cell bodies and dendrites of motoneurons innervating inspiratory muscles. Bulbospinal neurons were distributed throughout the dorsomedial and ventrolateral medulla, from the spinomedullary junction through about 0.8 mm rostral to the obex. Almost all of the 104 IR units recorded were located in corresponding parts of the ventrolateral medulla, rostral to nucleus retroambigualis, where expiratory related units are found. Injections of biotinylated dextran amine at the recording sites labeled projections both to the spinal cord and to the brainstem. In the lower brachial and upper thoracic spinal cord, bulbospinal axons traveled predominantly in the contralateral dorsolateral funiculus and terminated in close relation to the dendrites of inspiratory motoneurons retrogradely labeled with cholera toxin B-chain. In the brainstem, there were predominantly ipsilateral projections to the nucleus retroambigualis, tracheosyringeal motor nucleus (XIIts), ventrolateral nucleus of the rostral medulla, infraolivary superior nucleus, ventrolateral parabrachial nucleus, and dorsomedial nucleus of the intercollicular complex. In all these nuclei, except XIIts, retrogradely labeled neurons were also found, indicating reciprocity of the connections. These results suggest the possibility of monosynaptic connections between inspiratory premotor neurons and inspiratory motoneurons, which, together with connections of IR neurons with other brainstem respiratory-vocal nuclei, seem likely to mediate the close coordination that exists in birds between the vocal and respiratory systems. The distribution of IR neurons in birds is similar to that of the rostral ventral respiratory group (rVRG) in mammals.     

Anterograde transport of horseradish-peroxidase conjugated isolectin B4 from Griffonia simplicifolia I in spinal primary sensory neurons of the rat

Anterograde transport of the isolectin B4 from Griffonia simplicifolia I (B4) conjugated to horseradish peroxidase (HRP) was investigated in rat somatic and visceral primary sensory neurons at different spinal levels. Injection of B4-HRP into the L5 dorsal root ganglion (DRG) resulted in labelling in the sural nerve, but not in the gastrocnemius nerves. Free nerve endings and lanceolate-like nerve endings were labelled in the lateral hindpaw skin. Labelled fibres were also observed in the greater splanchnic nerve following B4-HRP injection into the T10-11 DRGs. Electron microscopic examination of the labelled nerves showed that B4-HRP labelled exclusively unmyelinated axons. In the spinal cord, labelling was observed in the superficial dorsal horn, and additionally, although much more sparse, in the medial and lateral collateral projections following injections into the T10-11 DRGs. The results suggest that B4-HRP should be a suitable anterograde tracer of unmyelinated cutaneous and splanchnic but not muscle primary afferent fibres.     

Segmental and propriospinal projection systems of frog lumbar interneurons

Spinal interneuronal networks have been implicated in the coordination of reflex behaviors and limb postures in the spinal frog. As a first step in defining these networks, retrograde transport of horseradish peroxidase (HRP) was used to examine the anatomical organization of interneuronal circuitry in the lumbar spinal cord of the frog. Following neuronal degeneration induced by spinal transection and section of the dorsal and ventral roots, HRP was placed at different locations in the spinal cord and the positions of labeled neuronal cell bodies plotted using a Eutectics Neuron Tracing System. We describe four spinal interneuronal systems, three with cell bodies located in the lumbar cord and one with descending projections to the lumbar cord. Interneurons with cell bodies located in the lumbar cord include: (1) Lumbar neurons projecting rostrally. Those projecting to thoracic segments tended to be located in the lateral and ventrolateral gray and in the lower two-thirds of the dorsal horn, with projections that were predominantly uncrossed. Those projecting to the brachial plexus and beyond were located in the dorsal part of the dorsal horn (uncrossed) and in the lateral, ventrolateral, and ventromedial gray (crossed). (2) Lumbar neurons with segmental projections within the lumbar cord. These neurons, which were by far the most numerous, had both uncrossed and crossed projections and were distributed throughout the dorsal, lateral, ventrolateral, and ventromedial gray matter. (3) Lumbar neurons projecting to the sacral cord. This population, which arose mainly from the dorsal horn and lateral or ventrolateral gray, was much smaller than in the other systems. Neuronal density of some of these populations of lumbar interneurons appeared to vary with rostrocaudal level. Finally, a population of neurons with cell bodies in the brachial and thoracic segments that projects to the lumbar cord is described. The most rostral of these neurons were multipolar cells with uncrossed projections, while those with crossed projections were confined almost exclusively to the ventral half of the cord. The distribution of spinal interneurons reported here will provide guidance for future studies of the role of interneuronal networks in the control of movements using the spinal frog as a model system.     

Neurotrophins support the development of diverse sensory axon morphologies

The initial outgrowth of peripheral axons in developing embryos is thought to occur independently of neurotrophins. However, the degree to which peripheral neurons can extend axons and elaborate axonal arborizations in the absence of these molecules has not been studied directly because of exquisite survival requirements for neurotrophins at early developmental stages. We show here that embryonic sensory neurons from BAX-deficient mice survived indefinitely in the absence of neurotrophins, even in highly dissociated cultures, allowing assessment of cell autonomous axon outgrowth. At embryonic day 11 (E11)-E13, stages of rapid axon growth toward targets in vivo, Bax-/- sensory neurons cultured without neurotrophins were almost invariably unipolar and extended only a rudimentary axon. Addition of neurotrophins caused outgrowth of a second axon and a marked, dose-dependent elongation of both processes. Surprisingly, morphological responses to individual neurotrophins differed substantially. Neurotrophin-3 (NT-3) supported striking terminal arborization of subsets of Bax-/- neurons, whereas NGF produced predominantly axon elongation in a different subset. We conclude that axon growth in vitro is neurotrophin dependent from the earliest stages of sensory neuron development. Furthermore, neurotrophins support the appearance of distinct axonal morphologies that characterize different sensory neuron subpopulations.     

Sprouting of primary afferent fibers after spinal cord transection in the rat

After spinal cord injury, hyper-reflexia can lead to episodic hypertension, muscle spasticity and urinary bladder dyssynergia. This condition may be caused by primary afferent fiber sprouting providing new input to partially denervated spinal interneurons, autonomic neurons and motor neurons. However, conflicting reports concerning afferent neurite sprouting after cord injury do not provide adequate information to associate sprouting with hyper-reflexia. Therefore, we studied the effect of mid-thoracic spinal cord transection on central projections of sensory neurons, quantified by area measurements. The area of myelinated afferent arbors, immunolabeled by cholera toxin B, was greater in laminae I-V in lumbar, but not thoracic cord, by one week after cord transection. Changes in small sensory neurons and their unmyelinated fibers, immunolabeled for calcitonin gene-related peptide, were assessed in the cord and in dorsal root ganglia. The area of calcitonin gene-related peptide-immunoreactive fibers in laminae III-V increased in all cord segments at two weeks after cord transection, but not at one week. Numbers of sensory neurons immunoreactive for calcitonin gene-related peptide were unchanged, suggesting that the increased area of immunoreactivity reflected sprouting rather than peptide up-regulation. Immunoreactive fibers in the lateral horn increased only above the lesion and in lumbar segments at two weeks after cord transection. They were not continuous with dorsal horn fibers, suggesting that they were not primary afferent fibers. Using the fluorescent tracer DiI to label afferent fibers, an increase in area could be seen in Clarke's nucleus caudal to the injury two weeks after transection. In conclusion, site- and time-dependent sprouting of myelinated and unmyelinated primary afferent fibers, and possibly interneurons, occurred after spinal cord transection. Afferent fiber sprouting did not reach autonomic or motor neurons directly, but may cause hyper-reflexia by increasing inputs to interneurons.     

Lysosomal changes in rat spinal ganglia neurons after prolonged treatment with cisplatin

We performed morphometric and ultrastructural studies to determine the morphological response of rat spinal ganglion sensory neurons to prolonged administration of cisplatin up to a total dose of 18 mg/kg. We quantitated the different types of lysosomal system (LS) bodies present (primary and secondary lysosomes, lipofuscin granules) as well as multivesicular bodies in treated and control animals. Five rats were examined per group. This ultrastructural study on cisplatin-induced changes in LS of spinal ganglia neurons shows that the total area and total number of LS structures are significantly increased by cisplatin treatment. The main specific changes were increase in number of small-size lysosomes and increase in number of polymeric lipofuscin granules. Other alterations observed were presence of nucleolar segregation, patches of neurofilaments and deposits of osmiophilic material in the perikaryon and axon hillock, all indicating that sensory neurons are a major target of cisplatin.     

Altered receptive fields and sensory modalities of rat VPL thalamic neurons during spinal strychnine-induced allodynia

Altered receptive fields and sensory modalities of rat VPL thalamic neurons during spinal strychnine-induced allodynia. J. Neurophysiol. 78: 2296-2308, 1997. Allodynia is an unpleasant sequela of neural injury or neuropathy that is characterized by the inappropriate perception of light tactile stimuli as pain. This condition may be modeled experimentally in animals by the intrathecal (i.t.) administration of strychnine, a glycine receptor antagonist. Thus after i.t. strychnine, otherwise innocuous tactile stimuli evoke behavioral and autonomic responses that normally are elicited only by noxious stimuli. The current study was undertaken to determine how i.t. strychnine alters the spinal processing of somatosensory input by examining the responses of neurons in the ventroposterolateral thalamic nucleus. Extracellular, single-unit recordings were conducted in the lateral thalamus of 19 urethan-anaesthetized, male, Wistar rats (342 +/- 44 g; mean +/- SD). Receptive fields and responses to noxious and innocuous cutaneous stimuli were determined for 19 units (1 per animal) before and immediately after i.t. strychnine (40 microgram). Eighteen of the animals developed allodynia as evidenced by the ability of otherwise innocuous brush or air jet stimuli to evoke cardiovascular and/or motor reflexes. All (3) of the nociceptive-specific units became responsive to brush stimulation after i.t. strychnine, and one became sensitive to brushing over an expanded receptive field. Expansion of the receptive field, as determined by brush stimulation, also was exhibited by all of the low-threshold mechanoreceptive units (14) and wide dynamic range units (2) after i.t. strychnine. The use of air jet stimuli at fixed cutaneous sites also provided evidence of receptive field expansion, because significant unit responses to air jet developed at 13 cutaneous sites (on 7 animals) where an identical stimulus was ineffective in evoking a unit response before i.t. strychnine. However, the magnitude of the unit response to cutaneous air jet stimulation was not changed at sites that already had been sensitive to this stimulus before i.t. strychnine. The onset of allodynia corresponded with the onset of the altered unit responses (i.e., lowered threshold/receptive field expansion) for the majority of animals (9), but the altered unit response either terminated concurrently with symptoms of allodynia (6) or, more frequently, outlasted the symptoms of allodynia (10) as the effects of strychnine declined. The present results demonstrate that the direct, receptor-mediated actions of strychnine on the spinal processing of sensory information are reflected by changes in the receptive fields and response properties of nociceptive and nonnociceptive thalamic neurons. These changes are consistent with the involvement of thalamocortical mechanisms in the expression of strychnine-induced allodynia and, moreover, suggest that i.t. strychnine also produces changes in innocuous tactile sensation.