Kappa opioid receptor density is consistent along the rostrocaudal axis of the female rat spinal cord

Kappa opioid receptors (KORs) were immunocytochemically localized at four different levels of the spinal cord of normally-cycling female rats in estrus or diestrus. KOR labeling was primarily observed in fine processes and a few neuronal cell bodies in the superficial dorsal horn and the dorsolateral funiculus. Quantitative light microscopic densitometry of the superficial dorsal horn revealed that there were no significant differences in KOR densities among spinal segments C1--C2, T2, T13--L1, and L6--S1 in either the estrus or diestrus phases. These results suggest that the potential for KOR-mediated antinociceptive responses is consistent along the rostrocaudal axis of the female rat spinal cord.     

Quantitative autoradiographic localization of [125I]neuropeptide Y receptor binding sites in rat spinal cord and the effects of neonatal capsaicin, dorsal rhizotomy and peripheral axotomy.

Using in vitro quantitative receptor autoradiography the present study reports on the distribution and possible changes of [125I]neuropeptide Y (NPY) binding sites in the rat spinal cord following neonatal capsaicin treatment, dorsal rhizotomy and sciatic nerve section. In control spinal cord the highest density of [125I]NPY binding sites was noticed in the superficial layers of the dorsal horn whereas low-to-moderate densities of [125I]NPY binding sites were detected in the deeper dorsal horn and in the ventral horn. In comparison with control rats, neonatally treated capsaicin rats showed a significant (P less than 0.001) bilateral decrease in [125I]NPY binding sites in the superficial layers of the dorsal horn. Unilateral dorsal rhizotomy and unilateral sciatic nerve section also exhibited a significant (P less than 0.05) depletion in [125I]NPY labeling in the superficial layers of the dorsal horn ipsilateral to the surgery. These results suggest that a certain proportion of [125I]NPY receptor sites is located on the primary afferent fibers of the superficial layers of the dorsal horn. This peptide thus could play an important role in the modulation of nociceptive transmission by acting directly on primary afferent terminals.     

Propriospinal afferent and efferent connections of the lateral and medial areas of the dorsal horn (laminae I-IV) in the rat lumbar spinal cord

The different subdivisions along the mediolateral extent of the superficial dorsal horn of the spinal cord are generally regarded as identical structures that execute the function of sensory information processing without any significant communication with other regions of the spinal gray matter. In contrast to this standing, here we endeavor to show that neural assemblies along the mediolateral extent of laminae I-IV cannot be regarded as identical structures. After injecting Phaseolus vulgaris leucoagglutinin and biotinylated dextran amine into various areas of the superficial dorsal horn (laminae I-IV) at the level of the lumbar spinal cord in rats, we have demonstrated that the medial and lateral areas of the superficial dorsal horn show the following distinct features in their propriospinal afferent and efferent connections: 1) A 300- to 400-microm-long section of the medial aspects of laminae I-IV projects to and receives afferent fibers from a three segment long compartment of the spinal dorsal gray matter, whereas the same length of the lateral aspects of laminae I-IV projects to and receives afferent fibers from the entire rostrocaudal extent of the lumbar spinal cord. 2) The medial aspects of laminae I-IV project extensively to the lateral areas of the superficial dorsal horn. In contrast to this, the lateral areas of laminae I-IV, with the exception of a few fibers at the segmental level, do not project back to the medial territories. 3) There is a substantial direct commissural connection between the lateral aspects of laminae I-IV on the two sides of the lumbar spinal cord. The medial part of laminae I-IV, however, does not establish any direct connection with the gray matter on the opposite side. 4) The lateral aspects of laminae I-IV appear to be the primary source of fibers projecting to the ipsi- and contralateral ventral horns and supraspinal brain centers. Projecting fibers arise from the medial subdivision of laminae I-IV in a substantially lower number. The findings indicate that the medial and lateral areas of the superficial spinal dorsal horn of rats may play different roles in sensory information processing.     

Expression of lectin binding in the superficial dorsal horn of the rat spinal cord during pre- and postnatal development.

The plant lectin Bandeiraea simplicifolia I-B4 binds to the soma and central terminals of a subpopulation of unmyelinated primary sensory neurones in the adult rat. The binding site of this lectin is thought to be the terminal alpha-D-galactose residue of a membrane associated glycoconjugate which may be involved in the development of specific connections between small diameter primary sensory neurones and second order neurones in the superficial dorsal horn of the spinal cord. To begin to investigate this possibility we have examined the development of lectin binding in the dorsal horn of pre- and postnatal rats. Lectin binding first appeared on axon profiles in the superficial dorsal horn of the spinal cord at embryonic days 18/19. Previous studies in the rat have revealed that the central processes of small diameter primary sensory neurones enter the dorsal horn at embryonic days 18/19. Our findings suggest that the glycoconjugate to which this lectin binds, is expressed by the central processes of small diameter primary sensory neurones as they grow into the spinal cord. It is therefore possible that this glycoconjugate is involved in the development of topographically ordered neural connections within the dorsal horn of the spinal cord.     

Galanin Receptor Binding Sites in Adult Rat Spinal Cord Respond Differentially to Neonatal Capsaicin, Dorsal Rhizotomy and Peripheral Axotomy

The discrete distribution and possible changes in specific [¹²⁵I]galanin binding sites were evaluated in the rat spinal cord following neonatal capsaicin treatment, dorsal rhizotomy and sciatic nerve section. The highest density of [¹²⁵I]galanin binding sites in the normal rat spinal cord was particularly evident in the superficial layers of the dorsal horn whereas moderate to low amounts of labelling were associated with the deeper dorsal horn, areas around the central canal and the ventral horn. Capsaicin-treated rats, compared to littermate controls, showed a significant bilateral increase in [¹²⁵I]galanin binding in the superficial laminae of the dorsal horn. Similarly, unilateral dorsal rhizotomy evoked a significant increase in the density of [¹²⁵I]galanin binding sites in the superficial dorsal horn ipsilateral to surgery. Section of the sciatic nerve, on the other hand, induced a significant depletion in [¹²⁵I]galanin binding in laminae I and II of the ipsilateral dorsal horn. These results, in parallel to those reported for galanin immunoreactivity under similar conditions, suggest that [¹²⁵I]galanin binding sites are preferentially located postsynaptically to the primary afferent fibre terminals in the dorsal horn of the spinal cord. Thus it seems that galanin, at the level of the dorsal spinal cord, regulates the processing of nociceptive information by acting on its own class of specific receptors located postsynaptically to primary sensory terminals.     

The estrous cycle affects pseudorabies virus (PRV) infection of the CNS

Previous work had suggested that mucosal immunity may be affected by the stage of the estrous cycle. Here, susceptibility to a neurotropic virus infection at different stages of the estrous cycle was assessed in a rodent model after direct injection of the virus into visceral organs. In the first two experiments, female Sprague-Dawley rats were infected with pseudorabies virus (PRV, Bartha's K-strain) by injection into either the cervix or the kidney after monitoring their estrous cycle. After either 4- or 5-day survival period post-infection, the rats were euthanized by transcardially perfusion and peripheral and central nervous system tissues were removed for immunocytochemical staining. The number of infected neurons was counted in various regions. Statistical analysis revealed that: (1) the number of infected cells in the sympathetic or parasympathetic ganglion, or the dorsal root ganglia was not affected regardless of the stage of the estrous cycle after cervix injection with PRV; (2) in contrast, the number of infected neurons in the spinal cord was affected significantly by the stage of the estrous cycle during viral infection of the cervix; (3) after kidney infection, the number of infected neurons found within the spinal cord or dorsal root ganglia varied significantly across the estrous cycle. In both cases, animals infected in proestrus or estrus had fewer infected neurons than animals infected in diestrus I or diestrus II (proestrous and estrous animals had less than 20% of infected cells found in diestrus I or diestrus II rats). In the third experiment, older, persistent estrous or persistent diestrous rats were infected by kidney injection and given a 4-day survival period, prior to virus isolation from lower thoracic spinal cord. Animals in persistent estrous had significantly less virus per gram of tissue than the persistent diestrous rats. These data suggest that the CNS of animals in proestrus or estrus is less susceptible to PRV infection compared to animals in either diestrus I or diestrus II. Because estrogen replacement therapy is known to restore some immune functions during reproductive ageing, it is speculated that plasma estrogen levels modulate the central nervous system's susceptibility to viral infections.     

Influence of estrous cycle on vaginocervical sensitivity: a fos-immunohistochemical study of lumbosacral spinal cord

Expression of c-fos in L₅–S₁ spinal segments in response to mechanical vaginocervical stimulation was investigated in both cycling and ovariectomized females. The aim of this paper was to verify the influence of estrous cycle on females genital tract sensitivity using immunodetection of a neural activity endogenous marker. The results indicate that lumbosacral spinal Fos-labeling was highly increased in vaginocervical stimulated rats relative to control, and labeled neurons were present more intensively in the dorsal horn in comparison to other spinal areas. Significant differences in Fos-labeling were observed according to the estrous cycle stage at which the stimulation was applied. In estrous females, the response was greater than that obtained at diestrous and much greater than the response of proestrous females. The spinal Fos-labeling of ovariectomized females is equivalent to that of diestrous females. These results give evidence that the vaginocervical induced expression of c-fos is modulated by cyclic changes in circulating sex hormones, whereas results observed in ovariectomized females indicate the likely involvement of other mechanisms independent of ovarian hormones.     

Colocalization of Oestrogen Receptor lmmunoreactivity and Preproenkephalin mRNA Expression to Neurons in the Superficial Laminae of the Spinal and Medullary Dorsal Horn of Rats

A double-labelling procedure combining immunohistochemical staining with in situ hybridization using a radiolabelled cRNA probe was employed to demonstrate oestrogen receptor-like immunoreactivity and preproenkephalin-A mRNA in the medullary and spinal dorsal horn of female rats. Both markers labelled large numbers of neurons in the substantia gelatinosa and its trigeminal homologue. Many of these neurons were double-labelled, displaying both oestrogen receptor-like- immunoreactivity and preproenkephalin-A mRNA; cell counts showed that 40–60% of the of the oestrogen receptor-like-immunoreactive cells in the superficial laminae also were labelled for preproenkephalin-A mRNA, and that 60–70% of the preproenkephalin-A mRNA-labelled neurons in the same laminae displayed oestrogen receptor-like immunoreactivity. Previous studies have shown that oestrogen receptors can bind to the promoter region of the preproenkephalin-A gene, and studies on the hypothalamus have demonstrated that oestrogen regulates enkephalin expression in select neuronal populations. The present results demonstrate that enkephalinergic neurons in the superficial dorsal horn contain oestrogen receptors and suggest that oestrogen may play an important role in the modulation of sensory and nociceptive processing in the lower medulla and spinal cord.     

Anatomical evidence for genetic differences in the innervation of the rat spinal cord by noradrenergic locus coeruleus neurons

Pontospinal noradrenergic neurons located in the A5, A6 (locus coeruleus, LC), and A7 cell groups are the major source of the noradrenergic innervation of the spinal cord. We have recently examined the specific terminations of these three cell groups in the spinal cord and found that the LC provides the major noradrenergic innervation of the ventral horn, while the A7 and A5 cell groups innervate the dorsal horn and intermediate zone, respectively. However, the results of similar experiments from another laboratory have shown that noradrenergic neurons in the locus coeruleus primarily innervate the dorsal horn, while the A5 and A7 innervate the intermediate zone and the ventral horn. These conflicting results may be due to fundamental genetic differences between the rats used in our experiments (Sasco Sprague-Dawley) and those used by the other laboratory (Harlan Sprague-Dawley). This possibility was examined by determining the projections of coeruleospinal neurons in these two rat substrains using the anterograde tracer Phaseolus vulgaris leucoagglutinin. The results indicate that in Sasco rats the LC neurons project through the ipsilateral ventromedial funiculus and terminate almost exclusively in the medial part of laminae VII and VIII, the motoneuron pool of lamina IX, and lamina X. In contrast, LC neurons in Harlan rats project bilaterally through the superficial dorsal horn and the dorsolateral funiculus and terminate most heavily in dorsal horn laminae I-IV. In addition, the LC neurons of Sasco rats innervate cervical spinal cord segments more densely than lumbar spinal cord segments, while in Harlan rats the lumbar spinal cord is more densely innervated than the cervical spinal cord. These results indicate that the projections of coeruleospinal neurons in Sasco rats are fundamentally different from those in Harlan rats and suggest that noradrenergic LC neurons may have different physiological functions in these two rat substrains.     

Glutamate uptake is attenuated in spinal deep dorsal and ventral horn in the rat spinal nerve ligation model

Alteration of glutamatergic (GLU) neurotransmission within the spinal cord contributes to hyperalgesic and allodynic responses following nerve injury. In particular, changes in expression and efficacy of glutamate transporters have been reported. Excitatory, pain transmitting primary afferent neurons utilizing glutamate as an excitatory neurotransmitter project to both superficial (I-II) and deep (III-V) laminae of the dorsal horn. These experiments were designed to examine changes in glutamate uptake occurring concomitantly within the spinal deep dorsal and ventral horn in situ after experimentally induced neuropathic pain. In vivo voltammetry, using microelectrode arrays configured for enzyme-based detection of GLU were employed. Sprague-Dawley rats had either sham surgery or tight ligation of L5 and L6 spinal nerves (SNL). Four to six weeks later, the L4-L6 spinal cord of chloral hydrate-anesthetized animals was exposed, and ceramic-based glutamate microelectrodes equipped with glass micropipettes 50 microm from the recording surfaces were placed stereotaxically at sites within the spinal cord. Pressure ejection of GLU into the ipsilateral L5-L6 spinal cord resulted in a 72% reduction of GLU uptake in SNL rats compared to sham controls in the ipsilateral L5-L6 deep dorsal horn and a 96% reduction in the ventral horn. In contrast, in the same animals, the contralateral L5-L6 or the ipsilateral L4 spinal cord showed no change in glutamate uptake. The data suggest that spinal nerve ligation produced attenuated glutamate uptake activity extending into the deep dorsal and ventral horn. The study suggests that plasticity related to spinal nerve injury produces widespread alteration in glutamate transporter function that may contribute to the pathophysiology of neuropathic pain.     

Arborization of single axons of the spinal dorsolateral funiculus to the contralateral superficial dorsal horn

This study provides an anatomical basis for the observation that a unilateral lesion of the spinal dorsolateral funiculus (DLF) can reduce the inhibitory effect of electrical stimulation of the nucleus raphe magnus (NRM) on dorsal horn nociceptive neurons located caudal and contralateral to the lesion. We injected the anterograde tracer Phaseolus vulgaris leucoagglutinin (PHA-L) into the NRM and traced the arborization of single DLF axons in the spinal gray matter. Although the majority of DLF axons arborized in the dorsal horn ipsilaterally, we found some axons which entered the spinal gray matter, traversed the gray matter to the central canal and then abruptly changed direction and coursed to the contralateral superficial dorsal horn. Very few branches were given off en route. These data indicate that some raphe-spinal axons may selectively influence the firing of neurons of the superficial dorsal horn, contralateral to the DLF in which they descend the spinal cord.     

Enkephalinergic neurons express 5-HT3 receptors in the spinal cord dorsal horn: single cell RT-PCR analysis.

The analgesic effect of activation of 5-HT3 receptors at the spinal cord is attenuated by the opioid antagonist naloxone. Enkephalin-immunoreactive neurons in the spinal cord superficial dorsal horn are innervated by 5-HT-immunoreactive fibers. This prompted us to examine whether enkephalinergic dorsal horn neurons express 5-HT3 receptors. Using the technique of single-cell RT-PCR adapted for small neurons in the superficial dorsal horn, methionine-enkephalin sequence-specific PCR products were observed in about half of the neurons studied. Furthermore, 5-HT3 receptor mRNA was detected in approximately 25% of enkephalinergic neurons. These observations suggest that at least part of the antinociception elicited by activation of 5-HT3 receptors at the spinal cord may involve enkephalinergic dorsal horn neurons.     

Regulation of neuropilin 1 by spinal cord injury in adult rats

Using RT-PCR, in situ hybridization, Western blotting, and immunofluorescence, we have analyzed the expression of neuropilin 1 (Np1) in two models of spinal cord injury (spinal cord hemisection and dorsal column crush) and following dorsal root rhizotomy in adult rats. Our results show that Np1 RNA and protein are up-regulated in the spinal cord after all these lesions but remain unaltered in the adjacent dorsal root ganglia. In control animals, Np1 levels in the spinal cord are low and appear to be localized mainly in blood vessels, motoneurons, and in the superficial layers of the dorsal horn. After DCC and rhizotomy, Np1 is expressed de novo around the injury and in the deafferentated dorsal horn, respectively, mainly by OX42-positive microglial cells. Both lesions affect the sensory projections, and interestingly a consistent increase of Np1 signal is additionally seen in the dorsal horn where these projections terminate. Unexpectedly, this increase is bilateral after unilateral rhizotomy.     

Spinal afferents to functionally distinct periaqueductal gray columns in the rat: An anterograde and retrograde tracing study

The segmental and laminar organization of spinal projections to the functionally distinct ventrolateral (vlPAG) and lateral periaqueductal gray (lPAG) columns was examined by using retrograde and anterograde tracing techniques. It was found 1) that spinal input to both vlPAG and lPAG columns arose predominantly from neurons in the upper cervical (C1–4) and sacral spinal cord; 2) that there was a topographical separation of vl-PAG projecting and lPAG-projecting neurons within the upper cervical spinal cord; but 3) that below spinal segment C4, vlPAG-projecting and lPAG-projecting spinal neurons were similarly distributed, predominantly within contralateral lamina I, the nucleus of the dorsolateral fasciculus (the lateral spinal nucleus) and the lateral (reticular) part of lamina V. Consistent with the retrograde results, the greatest density of anterograde label, within both the vlPAG and lPAG, was found after tracer injections made either in the superficial or deep dorsal horn of the upper cervical spinal cord. Tracer injections made within the thoraco-lumbar spinal cord revealed that the vlPAG column received a convergent input from both the superficial and deep dorsal horn. However, thoraco-lumbar input to the lPAG was found to arise uniquely from the superficial dorsal horn; whereas the deep dorsal horn was found to innervate the “juxta-aqueductal” PAG region rather than projecting to the IPAG. These findings suggest that similar to spino-parabrachial projections, spinal projections to the lPAG (and juxta-aqueductal PAG) are topographically organised, with distinct subgroups of spinal neurons projecting to specific lPAG or juxta-aqueductal PAG subregions. In contrast, the vlPAG receives a convergent spinal input which arises from the superficial and deep dorsal horn of cervical, thoracic, lumbar, and sacral spinal segments. J. Comp. Neurol. 385:207–229, 1997. © 1997 Wiley-Liss, Inc.     

On neuronal homology: a comparison of similar axons in Musca, Sarcophaga, and Drosophila (Diptera: Schizophora)

Transganglionic transport of horseradish peroxidase (HRP) was used to study the organization of the thoracic spinal nerve projection to the dorsal horn in rats. Labeling was found in the superficial dorsal horn 16-20 hours after application of HRP to the cut ends of various spinal nerve rami. Labeling was restricted to the outer part of the substantia gelatinosa at these stages. Longer survivals (25-48 hours) gave rise to labeling of the deep part of substantia gelatinosa and deeper parts of the dorsal horn as well. The dorsal ramus projected to the lateral third of the horn from half a segment rostral to half a segment caudal to the entry segment. The ventral ramus projected to the medial two-thirds of the horn from 1 1/2 segments rostral to half a segment caudal to the entry segment. The two branches of the ventral ramus that were examined projected to separate medial and lateral compartments for the entire ventral ramus. There was a distinct lateromedial shift of the projection found from rostral through caudal levels within the projection compartment for each nerve. The results indicate that the dorsal horn projection of thoracic spinal nerve branches is organized in longitudinal compartments which are arranged in a strictly somatotopic fashion.     

Analysis of the unmyelinated primary sensory neurone projection through the dorsal columns of the rat spinal cord using transganglionic transport of the plant lectin Bandeiraea simplicifolia I-isolectin B4

We have examined the projection of unmyelinated primary sensory neurones through the dorsal columns of the rat spinal cord using transganglionic transport of the plant lectin Bandeiraea simplicifolia I-isolectin B4. A small volume of the lectin was injected into the sciatic nerve of anaesthetised rats to label the central terminals of nociceptive primary sensory neurones. Following a survival period of 7 days, transverse and longitudinal sections of the superficial dorsal horn, dorsolateral funiculus and the dorsal columns from spinal segments L4 through to T13 were screened for lectin transport using light and electron microscopy. Longitudinal sections of the thoraco-lumbar region of spinal cord were also examined for lectin binding. Light and electron microscopy revealed transganglionically transported and bound lectin in the superficial dorsal horn and dorsolateral funiculus of the L3 and L4 segments of spinal cord. However, no lectin transport or binding was observed within the dorsal columns at any level of spinal cord examined. From these results, we suggest that the unmyelinated neurones within the dorsal columns do not express the binding site for BSI-B4 and, as such, may be responsible for visceral rather than cutaneous sensation. In line with the theories regarding a postsynaptic dorsal column pathway, these results suggest that nociceptors that bind BSI-B4 are not involved in a direct ascending projection through the dorsal columns.     

Spinal release of immunoreactive dynorphin A(1−8) with the development of peripheral inflammation in the rat

Microprobes bearing immobilised antibodies to dynorphin A₍₁₋₈₎ were used to study the basal and evoked release of this prodynorphin derived peptide in the spinal cord of urethane anaesthetised normal rats and those with a peripheral inflammation. In the absence of any active peripheral stimulus the antibody microprobes detected immunoreactive (ir)-dynorphin A₍₁₋₈₎ in two areas (lamina I and laminae IV–V) in the dorsal horn of the spinal cord of normal rats. With the development of unilateral ankle inflammation over 3 to 5 days following subcutaneous injections of Freund's complete adjuvant, a basal presence of ir-dynorphin A₍₁₋₈₎ was found in both the dorsal and ventral horn regions of both sides of the spinal cord. Lateral compression of the ankles of the normal animals did not release ir-dynorphin A₍₁₋₈₎ during the period of stimulation, but this neuropeptide was detected in increased amounts in the ventral horn following the stimulus. By contrast, compression of inflamed ankles produced elevated levels of ir-dynorphin A₍₁₋₈₎ during the period of stimulus application at three major sites in the ipsilateral spinal grey matter. The largest peak was in the deep dorsal horn/upper ventral horn (laminae VI–VII), with further sites of significant release in the mid dorsal horn (laminae II–V) and the lower ventral horn. The observation that ir-dynorphin A₍₁₋₈₎ is physiologically released in the ventral and deep dorsal in addition to the superficial dorsal horn of the rat suggests an involvement of dynorphins in several aspects of spinal function.     

A thyrotropin-releasing hormone-containing system in the rat dorsal horn separate from serotonin

In this study, we report the identification of a thyrotropin-releasing hormone (TRH)-containing system in the dorsal horn of the rat spinal cord. This system is distinct from the TRH and serotonin (5-hydroxytryptamine, 5-HT) cotransmitter supraspinal system that has projections to the intermediolateral (IML) and ventral columns. Spinal cord sections from untreated rats, and those treated with colchicine or 5,7-dihydroxytryptamine (5,7-DHT) were processed using peroxidase-antiperoxidase (PAP) immunocytochemistry with nickel intensification. Results of the 5,7-DHT treatment were verified by quantifying TRH and 5-HT by radioimmunoassay (RIA) and high performance liquid chromatography (HPLC), respectively. Prominent immunocytochemical staining for TRH in the dorsal horn was seen in varicose fibers mainly in lamina II and superficial lamina III of the dorsal horn of the spinal cord of control rats. A few fibers were seen ascending into lamina I. A moderate number of fibers that were immunoreactive for 5-HT were primarily in laminae I and II. The distribution of TRH- and 5-HT-containing neurites in the IML and the ventral horn agreed with previously published reports. Rats treated with colchicine showed many small round TRH immunoreactive cells that were limited to laminae II/III of the dorsal horn. TRH immunoreactivity in the dorsal horn and IML was resistant to the effects of the selective serotonin neurotoxin, 5,7-DHT, while the ventral horn was depleted of TRH staining. Serotonin was almost completely eliminated in all spinal cord laminae. Quantitative biochemical studies showed significant, but non-parallel reductions of TRH and 5-HT in cervical, thoracic and lumbar spinal cord. These studies demonstrate the existence of TRH-containing cell bodies and terminals in the dorsal horn of the rat spinal cord. These findings provide evidence that a TRH-containing system exists in the dorsal horn of the rat and that it is distinct from the descending medullary raphe system that contains 5-HT; suggest that a population of TRH-containing fibers that project to the IML may not contain 5-HT; and confirm previously published results that 5-HT and TRH coexist in terminals in the ventral horn of the spinal cord.     

Spinal projections of the locus coeruleus and the nucleus subcoeruleus in the Harlan and the Sasco Sprague-Dawley rat

The descending projections of the locus coeruleus (LC) and the nucleus subcoeruleus (SC) to the lumbar spinal cord were examined in rats from two vendors using retrograde transport of fluorescent latex beads. There was a vendor difference observed which agrees with previous findings. The differential dorsal horn and ventral horn projections of the Harlan and the Sasco Sprague-Dawley rats, reported by Fritschy and Grzanna, and Clark and Proudfit were confirmed. In the Harlan rat more cells were labeled in the LC following injections in the dorsal horn. In contrast, in the Sasco rat, more cells were labeled in the LC from injections in the ventral horn. Although, in all studies, the LC in rats from these vendors projected to some extent to both the dorsal and the ventral horn. A difference in labeling was noted also for the depth of placement of the tracer in the dorsal horn. When the site of injection was in the nucleus proprius, a predominately contralateral projection of the LC was noted. In contrast, when horseradish peroxidase (HRP) gel implants were placed to include the superficial laminae, the cells in the LC were labeled predominately ipsilaterally. The SC has a major projection to the dorsal horn in the Harlan rats while cells in the SC were predominately labeled following ventral horn injections in the Sasco rats. These cells send mostly ipsilateral projections to the dorsal and ventral horn of the spinal cord. Double labeled studies confirmed that 91% of LC and 86% of SC neurons projecting to the spinal cord were noradrenergic. The present results confirmed a difference in the descending catecholamine projections of rats purchased from different vendors. These strain differences may prove useful in studies of motor and sensory systems.     

Peptide coexistence in axon terminals within the superficial dorsal horn of the rat spinal cord.

The somata of primary sensory neurons have been shown to contain up to four (and possibly more) neuroactive peptides. Although each of these peptides has been separately located in axon terminals within the superficial dorsal horn of the spinal cord, it is not clear whether multiple peptide coexistence is also a feature of terminal varicosities. The aim of this study was to determine whether the peptides substance P (SP) and calcitonin gene-related peptide (CGRP), which are colocalized in the somata of a large number of primary sensory neurons, coexist in the central terminals of these neurons in the spinal cord. The protein A-gold technique of antigen localization was used to screen single boutons in laminae I and II of the rats spinal cord for SP- and CGRP-like immunoreactivity at the ultrastructural level. The results show that SP and CGRP are colocalized within a large number of synaptic boutons in the superficial dorsal horn. Furthermore, evidence was obtained to suggest that both SP and CGRP may be found in the same synaptic vesicle within these boutons. These findings indicate that both SP and CGRP may be coreleased from single terminals in the superficial dorsal horn. This is of considerable interest in view of the reported interaction between SP and CGRP in nociceptive behavioral responses in the rat.