A quantitative correlation of substance P-, calcitonin gene-related peptide- and cholecystokinin-like immunoreactivity with retrogradely labeled trigeminal ganglion cells innervating the eye

To evaluate the intraganglionic organization of ocular sensory neurons in the guinea pig, we studied the retrograde axonal transport from the eye to the trigeminal ganglion of cholera toxin B subunit and then applied immunohistochemistry for substance P, calcitonin gene-related peptide and cholecystokinin. Retrogradely labeled cells were observed only in the anteromedial portion of the ipsilateral ganglion. We observed no somatotopical organization to trigeminal neurons containing any of these three peptides, either for cells projecting to the eye or for the ganglion as a whole. The relative proportion of neurons immunoreactive for each of these three peptides was similar among the population of neurons retrogradely labeled with cholera toxin B and among the population of neurons without direct projections to the eye.     

Gene expression of histamine H1 receptor in guinea pig primary sensory neurons: a relationship between H1 receptor mRNA-expressing neurons and peptidergic neurons

Pharmacological studies have suggested that a subgroup of primary sensory neurons is responsive to histamine via the histamine H1 receptor. We addressed this issue using in situ hybridization histochemistry with a cRNA probe for the guinea pig H1 receptor gene. About 15% of the trigeminal and lumber dorsal root ganglion (DRG) neurons, but none of nodose ganglion neurons, were intensely labeled with this probe. The H1 receptor mRNA-positive neurons were exclusively small in size, and were demonstrated to give rise to unmyelinated fibers by ultrastructural analysis of isolectin B4-labeling. However, the H1 receptor mRNA-expressing DRG neurons were not immunoreactive to substance P (SP) and calcitonin gene-related peptide (CGRP). A marked increase in the number of mRNA-positive DRG neurons were observed 1-5 days after a crush injury of the sciatic nerve (3-4-fold of the control value). These neurons turned mRNA-positive after the nerve crush were also mainly small-sized. The mRNA signals were detected in many peptidergic (SP/CGRP) neurons, in contrast to the normal state. On the other hand, in the neurons which showed intense labeling in the normal condition, the mRNA signals were down-regulated. These results suggest that primary sensory neurons include two kinds of H1 receptor-expressing sensory neurons, one expressing H1 receptor mRNAs in the normal state and the other up-regulating the mRNAs following the peripheral nerve damage.     

Differential expression of the mRNA for the vanilloid receptor subtype 1 in cells of the adult rat dorsal root and nodose ganglia and its downregulation by axotomy.

Sensitivity to the pungent vanilloid, capsaicin, defines a subpopulation of primary sensory neurons that are mainly polymodal nociceptors. The recently cloned vanilloid receptor subtype 1 (VR1) is activated by capsaicin and noxious heat. Using combined in situ hybridization and histochemical methods, we have characterized in sensory ganglia the expression of VR1 mRNA. We show that this receptor is almost exclusively expressed by neurofilament-negative small- and medium-sized dorsal root ganglion cells. Within this population, VR1 mRNA is detected at widely varying levels in both the NGF receptor (trkA)-positive, peptide-producing cells that elicit neurogenic inflammation and the functionally less characterized glial cell line-derived neurotrophic factor-responsive cells that bind lectin Griffonia simplicifolia isolectin B4 (IB4). Cells without detectable levels of VR1 mRNA are found in both classes. A subpopulation of the IB4-binding cells that produce somatostatin has relatively low levels of VR1 mRNA. A previously uncharacterized population of very small cells that express the receptor tyrosine kinase (RET) and that do not label for trkA or IB4-binding has the highest relative levels of VR1 mRNA. The majority of small visceral sensory neurons of the nodose ganglion also express VR1 mRNA, in conjunction with the BDNF receptor trkB but not trkA. Axotomy results in the downregulation of VR1 mRNA in dorsal root ganglion cells. Our data emphasize the heterogeneity of VR1 mRNA expression by subclasses of small sensory neurons, and this may result in their differential sensitivity to chemical and noxious heat stimuli. Our results also indicate that peripherally derived trophic factors may regulate levels of VR1 mRNA.     

Morphology of ganglion cells which project to the dorsal lateral geniculate and superior colliculus in the ground squirrel

We wished to determine whether retinal ganglion cells that have axons terminating in the dorsal lateral geniculate and/or the superior colliculus have specific sizes of somata, comprising only part of the entire size range of ganglion cell somata. If so, then perhaps the specific functional types described by Michael might be associated with morphological types based on soma size. HRP was injected into either the superior colliculus (SC) or dorsal lateral geniculate nucleus (LGd) of thirteen-lined ground squirrels. Soma diameter of labeled ganglion cells was measured and the relation between cell size and frequency determined. After SC injections HRP-filled cells were mostly small and medium-sized. They ranged in diameter from 3 to 14 microns and the mean diameter of labeled neurons was 7.35 microns. Cells labeled after SC injections were often distributed as doublets or triplets in the retina. After LGD injections the majority of labeled cells were medium and large-sized. They ranged from 4 to 18 microns in diameter with a mean of 9.1 microns and were more regularly spaced within the retinal region of labeled cells. Thus, the present results provide reason to believe that functional classes of ganglion cells in ground squirrels may be correlated with particular morphological types.     

Estrogen receptor expression in lumbosacral dorsal root ganglion cells innervating the female rat urinary bladder

We have investigated whether bladder afferent neurons are likely to be targets for circulating estrogens by mapping estrogen receptor (ER) distribution in lumbosacral dorsal root ganglia (DRG) of adult female rats. Sensory neurons innervating either the detrusor or trigone regions were identified by application of fluorescent retrograde tracer dyes to the bladder wall. Labelled neurons were classified by their immunoreactivity for either type of ER (ERalpha or ERbeta) and further compared with subpopulations of neurons containing substance P, calcitonin gene-related peptide and vanilloid receptor (a marker of polymodal nociceptors). Both ER types were expressed in numerous sensory neurons of either upper lumbar (L1/L2) or lower lumbar/sacral (L6/S1) ganglia and there was almost complete coexpression of ERalpha and ERbeta. ER-positive neurons were mainly small-medium size (18-25-microm diameter), indicating that they may be nociceptors and/or supply visceral targets. Most bladder-projecting neurons expressed ERs and the majority of these also expressed neuropeptides or vanilloid receptor. Afferent neurons supplying detrusor and trigone regions had similar immunohistochemical features. About a third of the bladder-projecting neurons expressed both ER and vanilloid receptor, suggesting a mechanism by which estrogens could influence bladder pain. The prevalence of different chemical classes of ER-positive bladder-projecting neurons was reflected throughout the entire population of neurons in dorsal root ganglia of these spinal levels, suggesting that neurons supplying other pelvic visceral targets may have similar chemical profiles. These results suggest that many functional classes of sensory neurons innervating the lower urinary tract are likely to be targets for circulating estrogens, including many nociceptor neurons. The coexistence of ERalpha and ERbeta suggests a broad range of potential mechanisms by which estrogens may exert their genomic effects in this system.     

Nerve lesions and the generation of pain

This review addresses the issue of how axotomy of peripheral nerve fibers leads to pain and hyperalgesia. The point of axotomy (the nerve injury site), the dorsal root ganglia, and the dorsal horn of the spinal cord are candidate sites for generation of the pain signal that is likely to be critical for maintaining the neuropathic pain state. This review considers neuropathic pain from a "systems" perspective, tracing concepts of neuropathic pain from the work of Henry Head to the present. Surprisingly, the nerve injury site and the dorsal root ganglion belonging to a transected spinal nerve do not give rise to spontaneous activity in putative C-fiber nociceptors. The intact nociceptor belonging to adjacent uninjured spinal nerves, however, does acquire abnormal spontaneous activity and a chemical sensitivity to catechols. It is suggested that partially denervated tissues in the nerve, skin, and other locations may release substances that, in turn, sensitize the intact nociceptors. These abnormalities in the intact nociceptor, which arise in the context of Wallerian degeneration, probably play a role in creating or maintaining the abnormal pain state. These considerations probably also apply to the understanding of pain arising in other neuropathies. The findings relative to the "intact" nociceptor provide a rationale by which to understand how therapies distal to the nerve injury site may diminish pain.     

Isolation of an mRNA binding protein homologue that is expressed in nociceptors

The peripheral detection of painful stimuli requires the activation of small-diameter primary afferent neurons known as nociceptors. We have exploited two features of nociceptor biology, expression of the high affinity receptor for nerve growth factor (TrkA) and sensitivity to capsaicin, to isolate novel proteins using a differential display cloning scheme. A resulting approximately 4.3-kb cDNA was isolated and sequence analysis inferred a approximately 157-kDa protein containing a signal/mitochondrial targeting peptide sequence. Due to its molecular weight and significant amino acid identity with 'human leucine-rich protein 130'[leucine-rich pentatricopeptide motif containing (LRPPRC)], we termed the cDNA candidate leucine-rich protein 157 (rLRP157). Western blot analysis of HEK293 cells over-expressing the candidate cDNA showed a single protein product of similar size to that found in rat dorsal root ganglion as well as in other neuronal tissues and cell lines. Although expressed in a wide variety of tissues, in situ hybridization and immunohistochemistry in dorsal root ganglion revealed that rLRP157 expression was restricted to the small-diameter neurons. Sequence identity with previously characterized mRNA binding proteins and its subcellular localization in sensory neurons suggest that rLRP157 is associated with mitochondrial function. Moreover, the genetic basis of French-Canadian Leigh syndrome, which confers a loss of mitochondrial cytochrome c oxidase and is characterized by neurodegeneration, was recently mapped to a mutation in the LRPPRC gene. Taken together with its expression in small-diameter sensory neurons, we hypothesize that rLRP157, the rat orthologue of the human LRPPRC, may play a role in the modulation of peripheral pain transduction and serve as a novel marker for nociceptor subtypes.     

Localization of two calcium binding proteins, calbindin (28 kD) and parvalbumin (12 kD), in the vertebrate retina

We used immunocytochemistry to locate two calcium binding proteins, calbindin (CaB) and parvalbumin (PV), in the retina of goldfish, frog, chick, rat, guinea pig, dog, and man. The location of CaB depended on the type of dominant photoreceptor cells in birds and mammals. In cone-dominant retinas such as those of the chick, CaB-like immunoreactivity was found in the cones, cone bipolars, and ganglion cells. Amacrine cells 5-12 microns across were also labeled. In rod-dominant retinas, such as those of the rat, guinea pig, and dog, horizontal cells, small amacrine cells (about 6 microns across), and cells in the ganglion cell layer were labeled. In the human retina, which has both cones and rods in abundance, cones, cone bipolars, ganglion cells, horizontal cells, and small and large amacrine cells were labeled. In the frog and goldfish, the level of CaB-like immunoreactivity was low. In the frog, a few cones, amacrine cells, and cells in the ganglion cell layer were labeled. No immunoreactive structures were seen in the goldfish retina. PV-like immunoreactivity was found in chicks, rats, and dogs. No such immunoreactive structures were seen in the other species. In the chick, only amacrine cells were labeled. In the rat, amacrine cells and several displaced amacrine cells were labeled. In the dog, in addition to amacrine cells and displaced amacrine cells, horizontal cells were strongly labeled. Thus, PV-like immunoreactivity was found in those elements relating to the modulation of the main pathway of the visual transmission system.     

Species-specific expression of cholecystokinin messenger RNA in rodent dorsal root ganglia

The expression of cholecystokinin (CCK) messenger RNA (mRNA) was examined in dorsal root ganglia of rat and guinea pig using in situ hybridization histochemistry and RNA (Northern) blot hybridization with synthetic oligodeoxyribonucleotide (oligomer) probes. In guinea pig, CCK mRNA was detected in small and medium-sized neuronal perikarya comprising approximately 10-15% of the total dorsal root ganglia cell population. In contrast, in neurons of rat dorsal root ganglia, CCK mRNA was not detectable. Northern blot analyses revealed a single CCK mRNA species of expected size (0.8 kb) in guinea pig, but not rat, dorsal root ganglia. A 0.8 kb CCK mRNA was, however, detected in cortex of both rat and guinea pig. These data suggest that CCK is normally not synthesized in neurons of rat dorsal root ganglia and that there are species differences in CCK gene expression in mammalian sensory ganglia.     

Development of calretinin immunoreactivity in the mouse inner ear

Calretinin is a calcium-binding protein of the EF-hand family. It has been previously identified in particular cell types of adult guinea pig, rat, and chinchilla inner ear. Development of calretinin immunoreactivity in the mouse inner ear was investigated from embryonic day 13 (E13) to the adult stage. In the adult mouse vestibule, calretinin immunoreactivity was present in the same structures as described for the rat and guinea pig: the population of afferent fibers forming calyx units and small number of ganglion neurons. The earliest immunoreactivity was found at E17 in vestibular hair cells (VHCs), then, at E19, in afferent fibers entering the sensory epithelia and in rare ganglion neurons. At postnatal day 4 (P4), a few vestibular nerve fibers and ganglion neurons were reactive. From this stage until P14, immunoreactivity developed in the calyx units and disappeared from VHCs. At P14, immunostaining was adult-like. In the adult mouse cochlea, immunoreactivity was present in the same cell populations as described in the rat: the inner hair cells (IHCs) and most of Corti's ganglion neurons. Calretinin immunoreactivity appeared at E 19-P0 in IHCs and ganglion neurons of the basal turn. At P1, outer hair cells (OHCs) of the basal turn were positive. Calretinin immunoreactivity then appeared in IHCs, OHCs, and ganglion neurons of the medial turn, then of the apical turn. At P4, all IHCs and OHCs and most of the ganglion neurons were immunostained. Immunoreactivity gradually disappeared from the OHCs starting at P10 and, at P22, only IHCs and ganglion neurons were positive. The sequences of appearance of calretini, n were specific to each cell type of the inner ear and paralleled their respective maturation. Calretinin was transiently expressed in VHCs and OHCs. © 1994 Wiley-Liss, Inc.     

Excitation of marginal and substantia gelatinosa neurons in the primate spinal cord: indications of their place in dorsal horn functional organization.

Electrophysiological recordings were made from superficial parts of the spinal dorsal horn in monkeys, using dye-filled micropipette electrodes to permit iontophoretic marking of the recording sites for subsequent histological recovery. Focal field potentials and unitary activity evoked by dorsal root volleys including slowly-conducting components (both myelinated and unmyelinated) were found in the posteromarginal zone and the substantia gelatinosa (SG). Unitary potentials identified as being of the type recorded from cellular regions were separated into categories according to which group of slowly-conducting fibers and which kinds of cutaneous stimulation evoked the discharge. Recording locations for units excited by volleys in myelinated fibers conducting under 35 m/sec, by the types of skin stimulation activating either high-threshold mechanoreceptors (nociceptors) or cooling thermoreceptors, and giving no evidence of suprathreshold C-fiber excitation were centered on the posteromarginal zone. In contrast, recording loci for units exhibiting a strong C-fiber excitation and responses to cutaneous stimulation known to effectively excite C-fiber polymodal nociceptors or C-mechanoreceptors were centered in the SG. There appeared varying degrees of convergence of primary afferent input to the neuronal units, although most showed substantial specificity in their afferent excitation. On the bases of these results and consideration of existing morphological data, it is proposed that the marginal zone is a major synaptic termination region for the afferent fibers from high-threshold mechanoreceptors, cooling thermoreceptors, and perhaps other receptors with fine myelinated peripheral fibers. The SG, on the other hand, is suggested to be the terminal region for all types of unmyelinated primary afferent sensory neurons, and to have the complex role of integrating and distributing this input.     

Differences in extrinsic innervation patterns of the small intestine in the cattle and sheep

After oral challenge of the pathological prion protein, the pathogen was first detected in the distal ileum and then deposited in the brain. The present study aims determining the possible neuronal transport pathways from the small intestine to the brain in the cattle and sheep using a tracer protein. After horseradish peroxidase was injected into the wall in the duodenum of the calf and lamb and in the ileum of the lamb, the greater part of labeled neurons was detected in the celiac and cranial mesenteric ganglion complex. In the dorsal root ganglia T3 to L4 of both animals, some sensory neurons were always found to be labeled. Some parasympathetic preganglionic neurons were labeled in the dorsal motor nucleus of the vagus nerve after injections into the duodenum of the cattle and sheep, but extremely a small number of them were labeled after ileal application. The number of labeled sensory neurons in the nodose ganglion after duodenal injections of the sheep was much greater than that after duodenal application of the cattle. After ileal injections in the sheep, practically no labeled sensory neurons were found in the nodose ganglion. These results suggest that the pathological prion protein is mainly transported to the spinal cord and brain via the sympathetic nervous system and partially via the sensory neurons in the dorsal root ganglia. The vagus nerve seems to contribute to the transport of the pathogen not from the ileum, but from the duodenum.     

Isolectin B(4)-positive and -negative nociceptors are functionally distinct.

Small-diameter sensory neurons that are primarily nociceptors can be divided neurochemically into two populations: isolectin B(4) (IB(4))-positive nonpeptidergic neurons, and IB(4)-negative peptidergic neurons. It has been shown that IB(4)-positive neurons depend on glial-derived neurotrophic factor (GDNF), whereas IB(4)-negative neurons depend on NGF for survival during postnatal development (Molliver et al., 1997). Furthermore, these two populations of nociceptors terminate in distinct regions of the superficial spinal cord. To date, however, no evidence exists that indicates whether these two groups of nociceptors have distinct functional roles in the process of nociception (Snider and McMahon, 1998). To search for functional differences, we performed whole-cell voltage and current-clamp recordings on acutely isolated adult mouse dorsal root ganglion neurons that were labeled with fluorescent IB(4). We found that IB(4)-positive neurons have longer-duration action potentials, higher densities of TTX-resistant sodium currents, and smaller noxious heat-activated currents than IB(4)-negative neurons. Furthermore, we show that NGF, but not GDNF, directly increases the number of neurons that respond to noxious heat. The different electrophysiological properties expressed by IB(4)-positive and -negative small neurons, including their different heat sensitivities, indicates that they may relay distinct aspects of noxious stimuli both acutely and after injury in vivo.     

Histamine H1 receptor mRNA is expressed in capsaicin-insensitive sensory neurons with neuropeptide Y-immunoreactivity in guinea pigs

Histamine H1 receptor mRNA-expressing sensory neurons in guinea pigs are unmyelinated and are not immunoreactive to substance P and calcitonin gene-related peptide (CGRP) [Mol. Brain Res. 66 (1999) 24], which are implicated in the nociceptive transmission of the primary sensory system. In this study, we examined whether these H1 receptor mRNA-expressing neurons are sensitive to capsaicin by using in situ hybridization histochemistry. Of lumbar dorsal root ganglion (DRG) neurons in control animals, 17% were positive for CGRP. In guinea pigs neonatally treated with capsaicin (50 mg/kg), few CGRP-immunoreactive neurons were seen in the DRGs. However, the percentages of H1 receptor mRNA-expressing neurons (15-20%) and the intensity of the mRNA signals in these neurons were not affected by neonatal capsaicin treatment. We also revealed the presence of both capsaicin-sensitive and insensitive neuropeptide Y (NPY)-immunoreactive neurons in the DRGs. These neurons were exclusively small. H1 receptor mRNA was expressed in NPY-immunoreactive neurons in naive guinea pig DRGs. These results suggest that H1 receptor mRNA is expressed in capsaicin-insensitive DRG neurons with NPY-immunoreactivity in guinea pigs.     

Diversity of expression of the sensory neuron-specific TTX-resistant voltage-gated sodium ion channels SNS and SNS2

The differential distribution of two tetrodotoxin resistant (TTXr) voltage-gated sodium channels SNS (PN3) and SNS2 (NaN) in rat primary sensory neurons has been investigated. Both channels are sensory neuron specific with SNS2 restricted entirely to those small dorsal root ganglion (DRG) cells with unmyelinated axons (C-fibers). SNS, in contrast, is expressed both in small C-fiber DRG cells and in 10% of cells with myelinated axons (A-fibers). All SNS expressing A-fiber cells are Trk-A positive and many express the vanilloid-like receptor VRL1. About half of C-fiber DRG neurons express either SNS or SNS2, and in most, the channels are colocalized. SNS and SNS2 are found both in NGF-responsive and GDNF-responsive C-fibers and many of these cells also express the capsaicin receptor VR1. A very small proportion of small DRG cells express either only SNS or only SNS2. At least four different classes of A- and C-fiber DRG neurons exist, therefore, with respect to expression of these sodium channels.     

Genesis of neurons in the retinal ganglion cell layer of the monkey.

We have analyzed the genesis of various neuronal classes and subclasses in the ganglion cell layer of the primate retina. Neurons were classified according to their size and the time of their origin was determined by pulse labeling with 3H-thymidine administered to female monkeys 38 to 70 days pregnant. All offspring were sacrificed postnatally, and their retinas processed for autoradiography. The somata of cells in the retinal ganglion cell layer generated on embryonic day (E) 38 ranged from 9 to 14 microns in diameter. Between E40 and E56, the minimum soma diameter remained around 8-9 microns, while the maximum gradually increased to 22 microns. As a consequence, the means of the distributions of labeled cells also increased with age, from 11.8 microns diameter for cells generated on E38 to 14.6 microns diameter at E56. Over this period the percentage of labeled cells in the 10.5-16.5 microns and greater than 16.5 microns diameter range gradually increased. The proportion of the labeled cells in the less than 10.5 microns diameter range decreased from E38 to E45, but subsequently increased rapidly. At the end of neurogenesis in the retinal ganglion cell layer, around E70, most labeled cells were considerably smaller (7-9 microns) than those generated earlier. Our results indicate that within the ganglion cell layer of the macaque, neurons of small caliber are generated first, followed successively by medium sized cells. Large, putative P alpha cells are generated late. The production between E56 and E70 of cells with the smallest somata suggests that the last-generated neurons in the ganglion cell layer are predominantly displaced amacrine cells. Within the same sector of retina, different classes of neurons in the ganglion cell layer of the rhesus monkey appear to have a sequential schedule of production.     

Difference in binding by isolectin B4 to trkA and c-ret mRNA-expressing neurons in rat sensory ganglia.

The neurons labeled by isolectin B4 (IB4) in rat and mouse sensory ganglia are often regarded as non-nerve growth factor (NGF)-dependent and non-peptidergic neurons, but a considerable number of IB4-positive neurons in the dorsal root ganglion (DRG) are also shown to be immunoreactive to substance P (SP) and calcitonin gene-related peptide (CGRP), which are synthesized by NGF-dependent neurons. Therefore, we examined the relationships between the IB4-binding neurons and NGF/glial cell line-derived neurotrophic factor (GDNF)/GDNF-related proteins(GDNFs)-dependent neurons in rat DRGs by use of in situ hybridization histochemistry in serial sections. Of the DRG neurons, 42% and 22% were intensely and weakly labeled by IB4, respectively. The former neurons were small, and the latter varied in size. Of the trkA mRNA-expressing neurons, 29% and 57% were intensely and weakly labeled by IB4, respectively. On the other hand, 66% and 10% of the c-ret mRNA-expressing neurons were intensely and weakly labeled, respectively. The mRNA of somatostatin, another major neuropeptide in the sensory neurons, was exclusively expressed in the intensely IB4-labeled neurons. These findings suggest that many NGF-dependent and peptidergic sensory neurons are labeled by IB4 in rats.     

Primary sensory ganglion cells projecting to the principal trigeminal nucleus in the mallard, Anas platyrhynchos

The trigeminal and glossopharyngeal ganglia of the adult mallard were studied following HRP injections into the principal trigeminal nucleus (PrV). The PrV consists of the principal trigeminal nucleus proper (prV) and the principal glossopharyngeal nucleus (prIX). After an injection into the prV, the labeled cells were found in the ipsilateral trigeminal ganglion. After an injection into the prIX, labeled cells were found in the ipsilateral distal glossopharyngeal ganglion, but not in the proximal ganglion of the IX and X cranial nerve (pGIX + X). In Nissl preparations, two types of ganglion cells in the trigeminal ganglion, pGIX + X, and distal ganglion of N IX could be distinguished: larger light cells and smaller dark cells. We could not determine whether the HRP-labeled cells belonged to both types or to one of them; but because all the labeled cells were over 20 microns, we concluded that the smallest cells (10-19 microns) in the trigeminal ganglion and distal ganglion of N IX did not project to the PrV. The labeling of the cells in the distal ganglion of N IX (average 34.5 microns) was uniformly moderate. In the trigeminal ganglion there were two types of labeled cells: heavily labeled cells (average 29.1 microns) and moderately labeled cells (average 35.1 l microns). These two types of labeling (moderate and heavy) may reflect two types of primary sensory neurons: cells with ascending, nonbifurcating axons, and cells with bifurcating axons. We speculate that the former are proprioceptive neurons and the latter tactile neurons. Labeled bifurcating axons in the sensory trigeminal complex gave off collaterals to all parts of the descending trigeminal nucleus except to the caudalmost laminated spinal part.     

Compensatory projection of primary nociceptors and c-fos induction in the spinal dorsal horn following neonatal sciatic nerve lesion

The sciatic nerve was cut in newborn rats, and prevented from regenerating for 8 weeks. The number of dorsal root ganglion (DRG) neurons in L4 and L5, the distribution of central axon terminals of primary nociceptors, and the activity of secondary nociceptors were examined in the lumbar dorsal horn. The neonatal sciatic lesion caused about 60% reduction of DRG neurons. The central terminal field of the sciatic primary nociceptors negatively labeled by in situ binding of Bandeiraea simplicifolia isolectin B4 (BsIB4) markedly shriveled. Instead, the central representation of the saphenous nerve and the posterior cutaneous nerve of the thigh (PC) expanded. The laminae I/II neuropil in the medialmost (1/4) of the L3 dorsal horn and in the second lateral (1/4) around the L4/5 junction was occupied by the BsIB4 binding sites derived from the saphenous and the PC primary neurons, respectively. Noxious stimuli applied to the receptive fields of the saphenous and the PC nerves induced c-Fos-like immunoreactivity in many neurons in the expanded central terminal fields of the nerves. The collateral sprouts of uninjured primary nociceptors did not only invade the deafferented area of the dorsal horn but also established functional synaptic connections.     

Somatostatin-like immunoreactivity in primary afferents of the medial articular nerve and colocalization with substance P in the cat

The proportion of somatostatin-containing dorsal root ganglion cells innervating the knee joint of the cat via the medial articular nerve was determined by using retrograde labeling with fast blue and immunohistochemistry. Immunoreactivity was found in 8.6% of labeled cell bodies. In colchicine-treated ganglia, the proportion increased to 16.8%. Only small and intermediate-sized perikarya showed somatostatin-like immunoreactivity, indicating that this neuropeptide is synthesized predominantly in primary afferent units with unmyelinated sensory axons but may also be present in primary afferents with thinly myelinated sensory fibers. Colchicine treatment had no influence on the cell size distribution. Colocalization of somatostatin with substance P was determined by comparing the proportions of immunopositive dorsal root ganglion cells after incubation with antibodies against substance P or somatostatin or with a mixture of both. Substance P-like immunoreactivity was found in 18.1% (untreated ganglia) and 19.6% (colchicine treated ganglia) of the labeled neurons. After incubation with a mixed antibody solution, 18.2% of joint afferents in untreated and 19.9% of the cells in colchicine-treated ganglia were immunopositive. Comparing this result with the results obtained using somatostatin and substance P antibodies alone, one can calculate that both neuropeptides are colocalized in about 17% of the cat's knee joint afferents. About 3% of the neurons contain only substance P, whereas almost none of the neurons contain only somatostatin. Based on this fact, one can assume that both neuropeptides are coreleased in peripheral tissue as well as in the central nervous system. © 1995 Wiley-Liss, Inc.