Anatomical demonstration of gamma motoneurons in the cat's spinal cord

The modified Gudden method of Brodal was used in an attempt to localize anatomically the γ-motoneurons of the spinal gray matter. In thirty kittens, aged 3–12 days, various limb nerves were sectioned. The kittens were killed 2–10 days after operation and representative segments of the cord were fixed in alcohol, embedded in paraffin and subsequently cut in serial transverse sections and stained with thionine. Typically, small nerve cells with retrograde changes were found intermingled with similarly affected large α-motoneurons in Rexed's lamina IX. Since the γ-fibers of peripheral nerves are thin (< 8 μ), and based on the very likely assumption that small neurons have thin axons, it is concluded that the small neurons exhibiting retrograde changes in the present investigation were γ-motoneurons. They composed 10–15 per cent of all nerve cells in the spinal gray matter showing retrograde changes following section of limb nerves. Other small cells in lamina IX, which did not react to nerve section, were presumably interneurons.     

Rostrocaudal and laminar distribution of spinothalamic neurons in the high cervical spinal cord of the cat

In the cat, retrogradely labelled spinothalamic tract cells were found in contralateral medial laminae VI and lateral laminae VII-VIII and ipsilateral lateral laminae VII-VIII and medial laminae VI of the C1, C2, C3 and C4 segments. The proportion of cells in each ipsilateral laminae was similar in each segment but the proportion in contralateral medial laminae VI decreased while the proportion in contralateral laminae VII-VIII increased caudally.     

Morphological and physiological identification of excitatory pontine reticular neurons projecting to the cat abducens nucleus and spinal cord

The current study provides strong morphological and physiological evidence for identifying reticular neurons which project to the ipsilateral abducens nucleus. In conjunction with recent work in the alert cat, these neurons are believed to be excitatory and are implicated to play a role in the generation of saccadic and/or vestibular fast phase eye movements.     

Cortical neurons projecting to the cervical and lumbar enlargements of the spinal cord in young and adult rhesus monkeys

Cortical neurons projecting to cervical and lumbar segments of the spinal cord in five young and one adult monkeys were identified using the retrograde transport method following multiple unilateral injections of horseradish peroxidase (HRP) into the anterior horn at cervical and lumbar levels of the spinal cord. Somatotopically organized labeled neurons were found in the precentral and postcentral gyri, the rostral half of both the medial and dorsal aspects of area 5, the cingulate sulcus within the medial aspect of area 6, and the second somatosensory area within the lateral sulcus. All HRP-positive neurons were confined to cortical layer V or to a depth corresponding to the fifth layer in regions where delineation of cortical layers was obscured due to freezing or sectioning artifacts. Although cross-sectional areas of labeled neurons varied widely within each field, more large labeled neurons were present in the leg than in the arm subdivision of the precentral gyrus. HRP-positive neurons in the first and second somatosensory areas as well as those in the medial aspect of area 5 were of medium size, and those in the primary and supplementary motor areas as well as those within the dorsal aspect of area 5 were of medium or larger size.     

Evidence for periaqueductal cholecystokinin-substance P neurons projecting to the spinal cord

Using indirect immunofluorescence techniques combined with retrogradely transported fluorescent dyes, cholecystokinin (CCK)-like and substance P immunoreactive cell bodies in the periaqueductal central grey of the rat brain were studied. Data from both adjacent sections and elution-restaining techniques indicated that some of these central grey cells contain both a CCK-like peptide and substance P. Injection of the fluorescent dye, Fast Blue, into the cervical spinal cord indicated that this CCK-substance P cell group is a descending system. These findings provide evidence for two peptides in a single neuron, and the possible functional significance of such an occurrence is discussed.     

Sympathetic neurons in the cat spinal cord projecting to the stellate ganglion.

Horseradish peroxidase (HRP) was injected into the stellate ganglia of anesthetized cats. After a 2-day survival time the spinal cord was processed for the HRP reaction to study the distribution of the preganglionic neurons which project to the stellate ganglia. HRP-labeled neurons in the cord were located exclusively on the side ipsilateral to the injected stellate ganglia and were concentrated in four distinct areas: (1) intermediolateral cell column (78.2%), (2) lateral funiculus (18.5%), (3) intercalated area and (4) central autonomic area (3.3%) for the latter 2 areas). Their distribution pattern in the very rostral pole of the thoracic sympathetic preganglionic cell column was different from the typical thoracic pattern in that a greater proportion of the cells were located in the lateral funiculus, and a few labeled cells were observed in the ventral horn. Longitudinally, a wide range (C8-T8) of spinal cord levels projected to the stellate ganglia, with a peak at the T2 level. These observations infer that a large population of axonal processes of sympathetic preganglionic neurons traveled several segments of the spinal cord through an intraspinal pathway before exiting a particular ventral root.     

Anatomical features and physiological properties of central serotonin neurons

The authors describe the anatomical features and physiological properties of central serotonergic neurons. The central serotonin neurons (part of which store peptides [substance P, TRF, enkephalins] in addition to 5HT) are highly collateralized reticular-type brain stem neurons receiving multi-modal afferent information from ascending sensory and descending motor pathways. They are under control by noradrenergic, peptidergic and and gaba-ergic projection neurons and interneurons. Furthermore, they establish variable synaptoid and synaptic contacts to neuronal, glial and secretory targets throughout the entire neuraxis and send terminal branches into the ventricular CSF space. Firing rate and transmission activity appear to be controlled in a complex and rather rigid manner by 5HT release-dependent dendrodendritic and dendrosomatic inhibition via autoreceptors (which also regulate release at the axon terminals) and via transsynaptic inhibitory feedback circuits which may involve gabaergic projection and interneurons. 3H-imipramine appears to bind to an "imipramine recognition site" in the vicinity of the 5HT carrier, and to a variety of other transport and (postsynaptic) receptor sites (NA uptake, H1, 5HT2- and alpha 1-binding sites). Circumstantial evidence points to an as yet undetermined role of the postsynaptic 5HT-1-binding sites in neurotransmission. 5HT-2-binding sites fulfil the criteria for receptors: binding affinity of antagonists to these sites correlates significantly with their potency to inhibit behavioral excitation in rats elicited by 5-hydroxytryptophan or 5HT agonists.(ABSTRACT TRUNCATED AT 250 WORDS)     

Morphology and neurokinin 1 receptor expression of spinothalamic lamina I neurons in the rat spinal cord

Distinct morphological types of spinothalamic tract (STT) lamina I (LI) neurons have been identified in the cat and monkey spinal dorsal horn. Because these morphological types appear to differ in functional properties and receptor expression, we examined their distribution in the rat to test how their identification relates to earlier classification schemes. LI STT cells were retrogradely labeled with cholera toxin subunit b (CTb). Three types were recognized on the basis of cell body shape and proximal dendrites in the horizontal plane: fusiform, multipolar, and pyramidal. The relative distribution of these types was: 43, 26, and 28%, respectively, similar to that observed in the cat and monkey. 3D reconstructions were used to view each cell in all three major projection planes: horizontal, parasagittal, and transverse. Most LI STT neurons appeared fusiform in the parasagittal plane even though they belonged to different types based on their appearance in the horizontal plane, except in the most lateral portion of the dorsal horn, where LI curves ventrally. The proportion of STT neurons within LI was quantified by using the optical dissector method. To label all LI neurons, we used an anti-neuron-specific nuclear protein (NeuN) antibody. We found that approximately 9% of LI neurons projected to the thalamus. We also investigated neurokinin 1 receptor (NK-1r) expression in LI STT neurons. As in the monkey, most pyramidal STT neurons did not express NK-1r. These results provide further evidence that distinct morphological types of neurons differ in phenotype but not in their projection pattern.     

The location and function of respiratory fibres in the second cervical spinal cord segment: respiratory dysfunction syndrome after cervical cordotomy.

After high cervical percutaneous cordotomy for pain in malignant disease, 12 patients died during sleep at postoperative intervals between 1 and 8 days. Nine died after a first cordotomy and three after a second (contralateral) procedure. All except one had known pulmonary disease before operation. The operated segment of the spinal cord (C2) was studied histologically after death. Superposition of lesion outlines made it possible to determine those parts of the lesioned areas common to all unilateral and bilateral cases respectively. All cases dying of presumed respiratory dysfunction syndrome had lesions involving the region of the anterolateral funiculus in the C2 segment containing "pain" fibres activated from the second to fifth thoracic dermatomes. The fibres whose destruction appeared to be responsible for respiratory dysfunction syndrome were completely intermingled with ascending "pain" fibres. The possibility of these fibres being afferent in function is discussed.     

Anatomical organization of long ascending propriospinal neurons in the cat spinal cord

Retrograde transport of lectin-HRP conjugate (WGA-HRP) was used to examine the anatomical organization of long ascending propriospinal neurons (LAPNs) projecting to the cervical enlargement (C5-T1) and to the upper part of the cervical cord (C3-4) in cats. Small injections (0.05-1.0 microliter) of dilute (1-4%) WGA-HRP were made into the C5-T1 or C3-4 regions. The field potential evoked from stimulation of the superficial radial nerve served to position the micropipette delivering injections. Small and localized populations of labelled LAPNs were found in the dorsal horn (laminae IV-V), the intermediate zone (dorsal and medial lamina VII), and the ventral horn (ventral lamina VII, laminae VIII and IX). Ventral horn LAPNs projecting to the C5-T1 region were preferentially located in rostral lumbar regions. Ventral LAPNs projecting to the C3-4 region were more caudally situated. No regional differences in distribution of dorsal horn and intermediate zone LAPNs were noted in comparing the results of C3-4 with C5-T1 injection protocols. It is concluded that the caudally located ventral LAPNs may exert their influence on cervical motor output through C3-4 propriospinal interneurons. Other LAPNs are considered to exert their effect more directly, either at the C5-T1 or the C3-4 levels.     

A quantitative study of spinothalamic neurons in laminae I, III, and IV in lumbar and cervical segments of the rat spinal cord

The major ascending outputs from superficial spinal dorsal horn consist of projection neurons in lamina I, together with neurons in laminae III-IV that express the neurokinin 1 receptor (NK1r) and have dendrites that enter the superficial laminae. Some neurons in each of these populations belong to the spinothalamic tract, which conveys nociceptive information via the thalamus to cortical areas involved in pain. A projection from the cervical superficial dorsal horn to the posterior triangular nucleus (PoT) has recently been identified. PoT is at the caudal end of the thalamus and was not included in injection sites in many previous retrograde tracing studies. We have injected various tracers (cholera toxin B subunit, Fluoro-Gold, and fluorescent latex microspheres) into the thalamus to estimate the number of spinothalamic neurons in each of these two populations, and to investigate their projection targets. Most lamina I and lamina III/IV NK1r-immunoreactive spinothalamic neurons in cervical and lumbar segments could be labeled from injections centered on PoT. Our results suggest that there are 90 lamina I spinothalamic neurons per side in C7 and 15 in L4 and that some of those in C7 only project to PoT. We found that 85% of the lamina III/IV NK1r-immunoreactive neurons in C6 and 17% of those in L5 belong to the spinothalamic tract, and these apparently project exclusively to the caudal thalamus, including PoT. Because PoT projects to second somatosensory and insular cortices, our results suggest that these are major targets for information conveyed by both these populations of spinothalamic neurons.     

Postnatal development of brainstem serotonin-containing neurons projecting to lumbar spinal cord in rats

We quantified postnatal changes in brainstem serotonin (5-hydroxytryptamine, 5-HT)-containing neurons projecting to lumbar spinal cord. The medulla-spinal cord descending neurons were identified by a retrograde neurotracer, choleratoxin B subunit (CTb), and 5-HT neurons were stained by immunohistochemistry. Double-labeled neurons were assumed to be 5-HT neurons projecting to the lumbar spinal cord, and were quantitatively analyzed in each raphe nucleus in the medulla. The following results were obtained: (1) At PND 3, numerous CTb-labeled neurons (CTLN) were already present in the raphe pallidus (B1), while few CTLN were seen in raphe obscurus (B2) and raphe magnus (B3). CTLN then rapidly increased in number and were separately distributed after PND 7 in B3 and after PND 14 in B2. (2) At PND 3, numerous 5-HT-containing neurons were already present in B1-B3, with 23.4% and 14.0% of them labeled with CTb in B1 and B2, respectively, while there were few double-labeled neurons in B3. From PND 3 to 28, although the proportion of double-labeled to 5-HT neurons remained unchanged in B1 and B2, that in B3 rapidly increased from 5.8% at PND 7 to 28.8% at PND 14. Previous studies have shown that the 5-HT neurons in B3 send fibers mainly to the dorsal horn, while those in B1 and B2 send fibers mainly to the ventral horn at all spinal cord levels. Taken together, the present findings suggest that the brainstem 5-HT systems influence the ventral horn of the spinal cord, where spinal motoneurons exist earlier than in the dorsal horn. The functional significance of these early 5-HT systems in motor development and/or disabilities is discussed.     

Commissural neurons in the cat upper cervical spinal cord.

We have begun a study of the intrinsic circuitry of the cat's upper cervical cord, in part to elucidate the role of spinal interneurons in vestibulocollic reflexes. Using retrograde labelling with Fluoro-Gold and intraspinal microstimulation, we have identified commissural neurons projecting to the contralateral ventral horn. Neurons tended to be in the medial half of lamina VIII. Approximately half of the neurons were propriospinal neurons that could be activated antidromically from the rostral border of the cervical enlargement. Most of the tested, spontaneously active neurons were driven by stimulation of the ipsi- and/or contralateral vestibular nerve, in some cases disynaptically.     

Anatomical and physiological properties of pelvic ganglion neurons in female mice

Most neurons that regulate motility and blood flow in female pelvic organs are located within pelvic (paracervical) ganglia. In this study we investigated the anatomical and physiological properties of neurons within mouse (C57/Bl/6) paracervical ganglia. Most neurons showed immunoreactivity for choline acetyl transferase (CHAT) and were presumably cholinergic. Few neurons (approximately 5%) were tyrosine hydroxylase (TH) positive. Immunohistochemical labelling for microtubule associated protein 2 showed most neurons had small somata (cross sectional area approximately 300 microm(2)) and lacked dendrites. Action potential (AP) discharge characteristics, determined by depolarising current step injection, revealed most neurons (70%) adapted rapidly to depolarising current injection and were classified as "phasic". The remaining neurons discharged APs throughout the current step and were classified as "tonic". Membrane properties and current-voltage relationships were similar in phasic and tonic neurons, however the afterhyperpolarisation was significantly smaller in tonic neurons. Stimulation of preganglionic axons usually evoked a single strong preganglionic input (21/27 and 9/10 for pelvic and hypogastric nerves, respectively). In 19 preparations where we tested for inputs from both nerves pelvic inputs predominated (23/45 neurons) and inputs via the hypogastric nerve were rarely observed (3/45 neurons). Together, our data indicate that most neurons within mouse paracervical ganglia are cholinergic and parasympathetic. As there is little anatomical or functional evidence for integration of preganglionic inputs we propose that the role of paracervical neurons is restricted to one of spatial amplification or filtering of preganglionic inputs.     

Some properties of spinal neurons projecting to the medial brain-stem reticular formation

Experiments were done to locate and characterize neurons projecting from the spinal cord to the brain-stem reticular formation. In barbiturate-anesthetized or midcollicular-decerebrate cats, concentric bipolar electrodes were placed in the medial brain-stem reticular formation of the caudal pons and rostral medulla. Neurons were identified as projecting to brain-stem reticular formation using the antidromic collision criterion. Of 30 cells, 20 projected to the ipsilateral and ten to the contralateral brain-stem reticular formation. The mean conduction velocity was 49.6 m/sec. Cell somata were located deep to lamina V of the dorsal horn. Neurons in this sample fell into two categories based on their receptive field. One type of cell was excited by firm pressure over such deep structures as muscles, ligaments and periosteum. The second type had a large cutaneous receptive field. Neurons of this second type responded maximally to noxious cutaneous stimuli. Inhibitory receptive field components were common in neurons with both types of receptive field. In addition to antidromic activation, stimulation of medial brain-stem reticular formation produced both synaptic excitation and marked inhibition of some cells projecting to the brain-stem reticular formation. It is concluded that spinoreticular neurons may relay information concerning noxious stimuli.     

Response properties of spinal neurons projecting to midbrain or midbrain-thalamus in the monkey

Response and receptive field properties were determined for 24 spinal neurons backfired from midbrain or midbrain-thalamus in the anesthetized monkey. Recording sites were located in laminae I (11 cells), IV (6), V (6) and VI (1) in the lumbosacral spinal cord. The mean axonal conduction velocity of cells projecting to midbrain was significantly less than that of cells projecting to midbrain-thalamus. Cells backfired only from midbrain had complex excitatory receptive fields whereas those projecting to midbrain-thalamus had excitatory receptive fields confined to a single limb. Cells in both populations had complex inhibitory receptive fields.     

Anatomical and physiological studies of the gray matter surrounding the spinal cord central canal

Recent histochemical evidence suggests that neurons in the gray matter surrounding the central canal may play a role in nociception. We attempted to evaluate this possibility by studying the response properties and ascending projections of these cells in the rat. In the first series of experiments, the ascending projections of neurons around the central canal were studied by the method of retrograde transport of horseradish peroxidase (HRP). Predominantly contralateral labeling of neurons around the central canal appeared after HRP injections into the paramedian medullary or pontine reticular formation in intact or cerebellectomized animals. Far fewer cells were labeled by injections into the lateral medulla and/or pons. A small number of cells was retrogradely labeled by HRP injections into the vermal and intermediate regions of the cerebellum or the periaqueductal gray matter. Injections into other brainstem areas outside of the reticular formation also failed to label large numbers of neurons around the centra canal. In a second set of experiments, we recorded extracellular unitary activity from the lumbar enlargement in spinalized, decerebrate, unanesthetized rats. Thirteen units were functionally characterized and histologically localized within 300 micrometers of the central canal. All of the units identified responded exclusively to noxious stimuli applied within highly circumscribed receptive fields. Thus, neurons around the central canal contribute strongly to long ascending spinal cord projections. Physiologically, neurons found within this region are reminiscent of the noxious-specific cells in the outer most layers of the dorsal horn.     

Anatomical and physiological assessments of peripheral nerve grafts in the dorsal columns of the spinal cord

Portions of 1 cm length of the sensory radial nerve from the cat forelimb were used to replace an excised portion of the dorsal columns in the upper lumbar spinal cord. Observations were made on the clinical recovery of the animals, and cine recordings were made of their ability to traverse a horizontal ladder 5 months after the grafting procedure. Evoked sensory potential studies performed 6 months after grafting showed that an impulse arising from a stimulus applied to the sciatic nerve could be recorded in the spinal cord caudal to the graft, in the graft and in the spinal cord rostral to the graft in 5 out of 8 animals. Tracing of nerve connections with injection of horseradish peroxidase into the grafts resulted in labelling of nerve cell bodies in dorsal root ganglia and the grey matter of the lumbar spinal cord up to a distance of 10 mm away from the graft. These results confirm that peripheral nerve grafts can provide a satisfactory environment for the regrowth of ascending fibres in the dorsal columns of the spinal cord. However, there is as yet no evidence that the regenerated fibres succeed in forming useful synaptic connections with other nerve cell bodies.     

Segmental hypersensitivity and spinothalamic function in spinal cord injury pain

The mechanisms underlying central pain following spinal cord injury (SCI) are unsettled. The purpose of the present study was to examine differences in spinothalamic tract function below injury level and evoked pain in incomplete SCI patients with neuropathic pain below injury level (central pain) versus those without such pain. A clinical examination, quantitative sensory testing and magnetic resonance imaging (MRI) were performed in 10 SCI patients with below-level pain and in 11 SCI patients without neuropathic pain. Patients with and without pain had similar reductions of mechanical and thermal detection thresholds below injury level. SCI patients with central pain had sensory hypersensitivity in dermatomes corresponding to the lesion level more frequently than SCI patients without pain, but this may in part be explained by the exclusion of at-level spontaneous pain in the pain-free group. The rostral-caudal extent of the lesion measured by MRI did not differ between the two patient groups, and there were no statistically significant differences in any of the predefined areas of interest on the axial plane images. This study suggests that neuronal hyperexcitability plays a key role in central SCI pain and furthermore - in contrast to previous findings - that loss of spinothalamic functions does not appear to be a predictor for central neuropathic pain in spinal cord injury.