Lawrence Kruger

Diencephalic mechanisms of pain sensation

‘Universite Pierre et Marie Curie (Paris VI) and Groupe de Neurobiologie Appliqute, Laboratoire de Physiologie de la Nutrition, C. N. R. 7, 78350 Jouy en Josas (France); 2Department of Psychology, Florida State University, Tallahassee, FL, 32306 (U.S.A.); 3Departments of Anatomy and Anesthesiology and Ahmanson Laboratory of Neurobiology, Brain Research Institute, UCLA Center for the Health Sciences, Los Angeles, CA, 90024 (U.S.A.); “Department of Anatomy, School of Medicine, University of California, San Fransisco, CA, 94143; and *Departments of Physiology and Biophysics and Anatomy, Marine Biomedical Institute, University of Texas Medical Branch, Galveston, TX, 77550 (U.S.A.)

Selective neuronal glycoconjugate expression in sensory and autonomic ganglia: relation of lectin reactivity to peptide and enzyme markers

SummarySeveral plant lectins were used to characterize the cell-surface carbohydrates expressed on sensory ganglion cells and their central terminals in the spinal cord dorsal horn. In the rat, galactose-terminal glycoconjugates on a large subpopulation of small neurons whose central axons project to the substantia gelatinosa were demonstrated with the α-D-galactose-specificGriffonia Simplicifolia I-B4 (GSA) lectin. This neuron subset was labelled by alternative D-galactose-, N-Acetylgalactosamine-, and βGal(1,3)NAcGal-binding lectins. Similar GSA lectin reactivity was also illustrated in selected peripheral autonomic, gustatory and visceral sensory and enteric neurons, and the accessory olfactory bulb. The sensory neuron-specific isoenzyme, fluoride-resistant acid phosphatase (FRAP) co-localized with the GSA lectin, as did the monoclonal antibody (MAb) 2C5, which is directed against a lactoseries carbohydrate constituting a backbone structure of ABH human blood group antigens. In contrast, calcitonin gene-related peptide-immunoreactivity (CGRP-IR), used as a representative marker of peptidergic neurons, exhibited limited co-localization with GSA. A polyclonal anti-rat red blood cell (RBC) antibody co-localized with GSA, suggesting that lectin-reactive carbohydrates on rat sensory neurons are related to rat RBC antigens. In the human spinal cord, thel-fucose-bindingUlex europaeus-I (UEA) lectin also labelled the substantia gelatinosa; in rabbit, a small sensory ganglion cell subset and the spinal cord substantia gelatinosa was co-labelled by both the GSA and UEA lectins.These studies illustrate significant lectin-reactive cell surface carbohydrate expression by non-peptidergic, FRAP(+) sensory ganglion cells in the rat, and provide a means for visualizing the extensive, non-peptidergic, small sensory ganglion cell subpopulations, probably including a substantial proportion of nociceptive and unmyelinated peripheral axons.

Lectin and Neuropeptide Labeling of Separate Populations of Dorsal Root Ganglion Neurons and Associated “Nociceptor” Thin Axons in Rat Testis and Cornea Whole-Mount Preparations

As part of a program to explore patterns of innervation by nociceptor-related thin sensory axons in a variety of peripheral regions, we have labeled calcitonin gene-related peptide immunoreactive (CGRP-IR) nerve fibers in whole mounts of rat testicular tunica vasculosa and cornea. Efforts were undertaken to visualize the numerically significant fluoride-resistant acid phosphatase (FRAP)-containing axon population, whose peripheral endings have heretofore remained undemonstrable due to technical limitations of currently available acid phosphatase methods. Various histochemical markers that colocalize with FRAP in dorsal root ganglion (DRG) and spinal cord were examined, and a plant lectin, Griffonia simplicifolia I-B4, has been identified that not only selectively labels FRAP(+) sensory ganglion cells and central terminals in spinal cord, but also differentially stains a large number of thin axons in testicular and corneal whole mounts. Slender lectin-labeled fibers are abundant in cornea, and are distributed throughout tunica vasculosa preparations unrelated to blood vessels. CGRP-IR axons, in contrast, maintain close adherence to vascular patterns and are more coarse and varicose in appearance. Lectin staining therefore provides the first practical and specific method for visualization of peripheral FRAP(+) axons consisting principally of sensory C fibers but possibly including a small number of unmyelinated autonomic axons. It should now be feasible, using individual whole-mount preparations from various peripheral nociceptor-innervated tissues, to examine the distributions of both peptidergic and FRAP(+) fibers, which together comprise the vast majority of thin sensory axons. It may then be possible to correlate the observed anatomical patterns with knowledge regarding properties of corresponding physiologically characterized receptive fields.

Fixed-diameter polyethylene cuffs applied to the rat sciatic nerve induce a painful neuropathy: ultrastructural morphometric analysis of axonal alterations

&NA; Polyethylene cuffs of varying inner diameters were applied to the rat sciatic or sural nerve with the aim of inducing a standardized nerve injury, as assessed by morphometric analyses of fiber‐size spectrum alterations, associated with behavioral manifestations of neuropathic pain. The temporal sequence of axonal degeneration and regeneration was examined in parallel with behavioral analyses of pain initiation and recovery over a 6‐week postoperative (PO) period. Cuffs of 0.028–0.030″ inner diameter loosely enclosed sciatic nerves of young rats and elicited relatively uniform axonal degeneration and ‘pain’. Large myelinated axons underwent an early and sustained numerical depletion. Both the thinly myelinated and unmyelinated axon populations were initially diminished, but later rose to levels significantly greater than control values, likely the result of: (1) demyelination, (2) early stages of remyelination, (3) regenerative sprouting, and/or (4) collateral sprouting of undamaged unmyelinated axons. Pathological alterations of the injured nerve included edematous swelling, hypertrophy of the perineurial sheath, infiltration of fibroblasts and collagen into the intraneurial compartment, increasing interaxonal space and decreasing order and density of axonal packing. Animals displayed maximal pain‐related behaviors, including gait and postural asymmetries and hypersensitivity to mechanical compression and cold, during the 2nd week PO and had largely recovered by ∼ 4 weeks PO. Consistent behavioral manifestations of pain were achieved over a wide range of fiber spectrum alteration; however, with the largest cuffs or ‘bracelets’ used in this study, a substantial axonal fiber spectrum change was produced without inducing pain‐related behavior, suggesting that decrement in the number of myelinated axons was not always sufficient to elicit pain. Similar morphometric and pathological results were achieved with sural neuropathy after 0.010″ ID cuffs and 14 days PO survival. Considering the lack of correlation between axonal alterations and pain, modification in the local intraneurial microenvironment at the site of injury may be a key component of peripheral pain mechanisms; these include changes in the biochemical milieu, increased intraneurial pressure, and altered nociceptor sensitivity or impulse propagation in the relatively intact unmyelinated axon population.

Topography of the retinal projection upon the superior colliculus of the cat.

Abstract The distribution of receptive field locations in the visual field was determined for the entire surface of the superior colliculus of anesthetized cats by recording impulse discharges in the superficial layers with microelectrodes. A reconstruction of the visual projection reveals an alinear binocular topography with marked expansion of the representation of the area centralis. At the rostral pole, a small zone displays a separate contralateral projection from the temporla retina (nasal field) containing monocularly excited units with distinctive properties. For the remainder of the colliculus. homonymous points in the visual field for each hemiretina overlap, with a tendency for contralateral dominance. A bilateral projection of the retinal area concerned with central vision may be related to the evolutionary acquisition of an uncrossed optic pathway.

An interpretation of dental innervation based upon the pattern of calcitonin gene-related peptide (CGRP)-immunoreactive thin sensory axons.

Calcitonin gene-related peptide (CGRP) is a recently characterized neuroactive substance that is expressed in a large proportion of small- to medium-diameter sensory ganglion neurons whose central terminals lie in the superficial spinal and medullary dorsal horn. This restricted distribution within the peripheral nervous system suggests a prominent role for the peptide in nociceptive processing. The mammalian tooth pulp, which receives a relatively homogeneous afferent input from thin (putative nociceptive) fibers originating from this subpopulation of trigeminal ganglion cells, thus affords an ideal target zone in which to examine peripheral nociceptive mechanisms. The large percentage of these neurons displaying CGRP-like immunoreactivity (CGRP-LI) furthermore provides a valuable tool to study its thin-fiber afferent innervation. CGRP-LI has been localized within intact, decalcified specimens of rat, cat, monkey, and human teeth and associated dental structures. A remarkably robust CGRP-LI innervation of molar pulp and dentin was revealed in all species, with fibers coursing both in fascicles and individually, in variable relation to blood vessels and pulpal stroma. Our methods enabled tracing of a large number of axons through Raschkows plexus and odontoblast layer into dentinal tubules. Paralleling anterograde axonal transport studies, a greater share of fibers was found in coronal vis-à-vis radicular dentin. In the rat, this fiber pattern stood in contrast both to incisor dentin, which appeared devoid of CGRP-LI, and to the abundant labeled axons in gingiva and periodontal tissues. Surgical deafferentation of rat mandible resulted in widespread depletion of CGRP-LI, while superior cervical ganglionectomy was without effect, confirming the sensory nature of the CGRP-LI fibers. Neonatal capsaicin treatment greatly attenuated the immunostaining, providing evidence for CGRP-LI localization in chemosensitive unmyelinated afferents. The great density of CGRP-LI axons demonstrated is considered in contrast to the restricted range and extent of sensory stimuli to which teeth are presumably subjected, and in relation to the diverse ongoing trophic, regulatory, and reparative processes in tooth structures. It is therefore suggested that these fibers may be subserving prominent efferent roles in dental pulp not directly related to nociception.

Deafferentation in animals as a model for the study of pain: an alternative hypothesis

The notion that post-deafferentation autonomy is a pain response is unsupported by the results of studies with neurotoxins. The selective massive destruction of a fiber system considered essential to normal nociception--unmyelinated primary afferent axons--prior to deafferenting nerve lesions did not stop or even significantly impede post-denervation DI despite massive evidence from humans and animals that pain following nerve lesions originates in the periphery and is generated by abnormal discharges in the injured nerve. In addition, when a reduction in abnormal impulse discharges of both large and small injured sensory axons could be inferred following neonatal sympathectomy, DI was not reduced in incidence or severity. This latter observation (1) provides further support for a dissociation between DI and pain, since any contribution of myelinated primary afferent axons to painful pathology probably was substantially reduced by sympathectomy and (2) suggests that DI also may be unrelated to non-painful sensory pathology attributable to abnormal activity in the thick-diameter fiber population. These findings and an evaluation of other relevant observations suggest that DI may not be a manifestation of deafferentation pain and perhaps this animal model for the experimental study of pain should be discarded. An alternative view of DI, reconcilable with known properties of this behavior, is that it reflects a proclivity in some species and circumstances to shed a functionally-impaired insensate appendage.

A morphological and somatotopic analysis of single unit activity in the trigeminal sensory complex of the cat.

Abstract The receptive fields of 788 neurons have been studied in order to reconstruct the relation of the spatial projection of the integument of the face onto different portions of the sensory trigeminal nuclear complex. At all rostrocaudal levels extending from the principal sensory nucleus to the upper cervical cord, the face is represented with the mandibular division lying dorsally, the ophthalmic division at the ventral surface and the interior of the buccal cavity in the medial portion. Thus each point on the face is represented by a colum of cells extending from the level of the principal V nucleus to at least the lower medulla or upper cervical cord. For a given zone of innervation, neurons within these columns can be excited by delicate tactile stimuli applied to receptive fields of a relatively narrow size range. The trigeminal tactile region of the brain stem conforms within the limits of the experimental method to the morphologically defined trigeminal sensory complex except for some findings beyond its medial limits, as usually defined, concerned with buccal cavity innervation. A revision of the subdivision of the architectonic units of the trigeminal complex is suggested on the basis of morphological and functional evidence.

Localization of specific binding sites for atrial natriuretic factor in peripheral tissues of the guinea pig, rat, and human.

Specific, high affinity atrial natriuretic factor (ANF) binding sites were identified and localized by autoradiographic techniques in peripheral tissues of the guinea pig, rat, and human. In the guinea pig kidney, high concentrations of ANF binding sites were located in the glomerular apparatus, outer medulla, and small renal arteries. Other peripheral tissues containing ANF binding sites included the zona glomerulosa of the adrenal cortex, the smooth muscle layer of the aorta and gallbladder, the lung parenchyma, the posterior lobe of the pituitary, the ciliary body of the eye, and the leptomeninges and choroid plexus of the brain. The distribution of ANF binding sites in the rat and human kidney was nearly identical to those seen in the guinea pig kidney; high concentrations were present in the glomerular apparatus, outer medulla, and small renal arteries. These results are consistent with earlier physiological and pharmacological studies that suggested that ANF plays a functional role in the regulation of extracellular fluid volume and blood pressure. There appears to be little species variation in the location and concentration of renal ANF binding sites, suggesting that, at least in the kidney, the results in experimental animals are relevant to the actions of ANF in humans. The finding that ANF binding sites were stable and present in high concentrations in human postmortem kidneys further suggests that these tissues may be amenable to testing for the involvement of ANF receptor dysfunction in diseases such as hypertension and congestive heart failure.

Tactile neurons of the superior colliculus of the cat: Input and physiological properties

Neurons in the strata intermediale and profundum of the cat superior colliculus (SC) previously have been shown to be organized somatotopically and to respond to tactile stimuli. The stimulus properties required for neuronal activation suggest that these neurons are excited by inputs from selected classes of high-velocity, sensitive cutaneous mechanoreceptors, and that stimulus-response relationships indicate only a narrow range of information concerning stimulus magnitude. The properties of these neurons have been interpreted as serving a role in stimulus detection that recovers slowly and may be best suited for directed attention and orientation behavior. The input to the SC was studied by injection of horseradish peroxidase (HRP) for retrograde labeling of neurons projecting to this region. The main inputs subserving the tactile pathway are the principal trigeminal nucleus and the rostral portion of the spinal V subnucleus oralis and the main cuneate nucleus. Sparser labeling was found in other somatic relays, including the gracile and lateral cervical nuclei and posterior portion of the spinal trigeminal complex. Several other projections relating to visual mechanisms are also described.