Brain-derived TNFalpha mediates neuropathic pain.

Neuropathic pain is a chronic pain state that develops a central component following acute nerve injury. However, the pathogenic mechanisms involved in the expression of this central component are not completely understood. We have investigated the role of brain-associated TNF in the evolution of hyperalgesia in the chronic constriction injury (CCI) model of neuropathic pain. Thermal nociceptive threshold has been assessed in rats (male, Sprague-Dawley) that have undergone loose, chromic gut ligature placement around the sciatic nerve. Total levels of TNF in regions of the brain, spinal cord and plasma have been assayed (WEHI-13VAR bioassay). Bioactive TNF levels are elevated in the hippocampus. During the period of injury, hippocampal noradrenergic neurotransmission demonstrates a decrease in stimulated norepinephrine (NE) release, concomitant with elevated hippocampal TNF levels. Continuous intracerebroventricular (i.c.v.) microinfusion of TNF-antibodies (Abs) starting at four days, but not six days, following ligature placement completely abolishes the hyperalgesic response characteristic of this model, as assessed by the 58 degrees C hot-plate test. Antibody infusion does not decrease spinal cord or plasma levels of TNF. Continuous i.c.v. microinfusion of rrTNF alpha exacerbates the hyperalgesic response by ligatured animals, and induces a hyperalgesic response in animals not receiving ligatures. Likewise, field-stimulated hippocampal adrenergic neurotransmission is decreased upon continuous i.c.v. microinfusion of TNF. These results indicate an important role of brain-derived TNF, both in the pathology of neuropathic pain, as well as in fundamental pain perception.

Serotonergic binding in the rat dorsal raphe nucleus: critical role of MAO inhibition.

The biochemical properties of the 5-HT1A receptor in dorsal raphe nucleus (DRN) were investigated using a micropunch procedure. Initially, the Ki value for 5-hydroxytryptamine (5-HT) binding to a site labeled by the 5-HT1A-selective ligand [3H]8-hydroxy-2-(di-n-propylamino)tetralin (8-OH-DPAT) was 20-fold higher than the KD for [3H]5-HT. In addition, a number of putative 5-HT1A selective ligands displayed poor affinity for the [3H]8-OH-DPAT site. The possibility that these discrepant results were due to metabolism of the receptor ligands was investigated by increasing the concentration of the monoamine oxidase (MAO) inhibitor, pargyline. Increasing the concentration of pargyline reduced, but did not abolish, the discrepancy between the Ki and KD values for 5-HT. However, inclusion of clorgyline, which is a more potent MAO inhibitor, resulted in an-excellent agreement between the Ki and KD values for 5-HT. In addition, when clorgyline was used, 5-HT1A-selective compounds displayed high affinity for the DRN binding site consistent with [3H]8-OH-DPAT labeling a 5-HT1A receptor in this tissue. The present study describes a fast and easy method for measuring biochemical properties in small discrete brain areas. These studies also indicate that pargyline should be replaced in serotonergic binding assays with a more potent inhibitor of monoamine oxidase such as clorgyline.

Effect of high-energy phosphates and free radical scavengers on replant survival in an ischemic extremity model.

In replantation surgery, preoperative and intraoperative ischemia can lead to irreversible changes that prevent reperfusion during the subsequent re-establishment of circulation. These changes are termed the no-reflow phenomenon. Ischemic phase damage was addressed by comparing the dose-response effects of controls vs. five different high-energy phosphate compounds on replanted limb survival. Reperfusion damage was evaluated via comparisons of controls with superoxide dismutase (SOD). Ischemic hindlimbs treated with high-energy phosphates displayed improved survival compared with controls. Limbs treated with SOD demonstrated no change in survival at 4 hours and improved survival at 8 hours. Combining adenosine and SOD had no improved effect on survival. Adenosine was the most effective high-energy phosphate in limiting ischemic damage. The free radical scavenger (SOD) was beneficial only at the later stages of ischemia. In this experimental model, there appears to be a role for both phosphates and free radical scavengers in enhancing ischemic tissue survival.

Co-cultures of microglia and astrocytes from kainic acid-lesioned adult rat hippocampus: effects of glutamate.

The long-standing question concerning the direct actions of glutamate on the membrane potential of astroglial cells in the central nervous system was addressed using the in vitro kainic acid-lesioned hippocampal slice preparation and primary cell co-cultures of astrocytes and microglia derived from such lesions. The ultrastructure of the lesioned hippocampus was examined to aid in the identification of the cells appearing in culture. In culture, microglia appeared as flat cells, less than 1 micron in thickness at the edge of the cell, but thicker (about 5 microns) near the nucleus. The cytoplasm was packed with granular inclusions. Microglia appeared in two morphological forms, amoeboid and ramified. The amoeboid form was characterized by a cell body with a single process, and was always observed 1 day after starting the cell culture. Such cells became less frequent after 1 week in culture. The ramified form appeared as a rounded cell, devoid of processes, and were frequently observed in older cultures (greater than 1 week). Microglia did not round up after exposure to dibutyrylcyclic adenosine monophosphate (cAMP), and did not stain for glial fibrillary acidic protein (GFAP). An ultrastructural examination of the lesion demonstrated that microglia were present and that they contained many cytoplasmic granules similar to lipofuscin-containing granules. No filaments were observed in the cytoplasm of microglia. By contrast, the cytoplasm of astrocytes in culture had far fewer granules, rounded up to dibutyryl-cAMP, exhibited multiple processes, and stained for GFAP. In slices, astrocytes had no lipofuscin-containing granules, but numerous cytoplasmic filaments were present.(ABSTRACT TRUNCATED AT 250 WORDS)

MR imaging of normal rat brain at 0.35 T and correlated histology.

A custom-built small-animal transceiver was used for in vivo imaging of normal rat brain at 0.35 T, with the objective of identifying anatomic components by comparison of images with corresponding histologic sections. The cerebrum, cerebellum, brain stem, ventricles, hippocampus, and subarachnoid space were identified and cerebrospinal fluid (CSF) was differentiated from gray matter and white matter on coronal and transaxial magnetic resonance (MR) images. These images compare favorably with those obtained by others at higher field strengths in regard to delineating major neuroanatomic structures. It is concluded that this technique will be useful for investigating small-animal models of human neurologic disease involving morphologic and morphometric changes in gray matter, white matter, and CSF-filled spaces.

Morphometry of the lumbar spine: anatomical perspectives related to transpedicular fixation.

The pedicles of lumbar vertebrae were measured both directly and radiographically to determine the differences between the sexes and the accuracy of radiographic measurement. The lumbar pedicles of cadavera of forty-nine patients--twenty-four men and twenty-five women--who died between the ages of sixty and ninety-eight years were measured directly and on radiographs. The pedicles of lumbar vertebrae from fifty-one patients--twenty-three men and twenty-eight women--between the ages of twenty and fifty years who had low-back problems were measured on radiographs and computerized tomographic scans. Comparison revealed that the average transverse and sagittal diameters of the pedicles and the distance from the posterior aspect of the laminar cortex to the anterior aspect of the cortex of the vertebral body along the central axis of the pedicles were 5 to 20 per cent greater in men, but the transverse and sagittal angles of the pedicle did not differ significantly between the sexes. Measurements on radiographs and computerized tomographic scans of the transverse angles of the pedicles and of the distances from the posterior aspect of the laminar cortex to the anterior aspect of the cortex of the vertebral body from the second to the fifth lumbar vertebra were greater than direct measurements, even without magnification. Direct measurements of the diameters of the transverse and sagittal diameters of the pedicle of the fifth lumbar vertebra, however, were greater than the radiographic measurements.

Afferent projections to the deep mesencephalic nucleus in the rat.

Afferent projections to the deep mesencephalic nucleus (DMN) of the rat were demonstrated with axonal transport techniques. Potential sources for projections to the DMN were first identified by injecting the nucleus with HRP and examining the cervical spinal cord, brain stem, and cortex for retrogradely labeled neurons. Areas consistently labeled were then injected with a tritiated radioisotope, the tissue processed for autoradiography, and the DMN examined for anterograde labeling. Afferent projections to the medial and/or lateral parts of the DMN were found to originate from a number of spinal, bulbar, and cortical centers. Rostral brain centers projecting to both medial and lateral parts of the DMN include the ipsilateral motor and somatosensory cortex, the entopeduncular nucleus, and zona incerta. at the level of the midbrain, the ipsilateral substantia nigra and contralateral DMN likewise project to the DMN. Furthermore, the ipsilateral superior colliculus projects to the DMN, involving mainly the lateral part of the nucleus. Afferents from caudal centers include bilateral projections from the sensory nucleus of the trigeminal complex and the nucleus medulla oblongata centralis, as well as from the contralateral dentate nucleus. The projections from the trigeminal complex and nucleus medullae oblongatae centralis terminate in the intermediate and medial parts of the DMN, whereas projections from the contralateral dentate nucleus terminate mainly in its lateral part. In general, the afferent connections of the DMN arise from diverse areas of the brain. Although most of these projections distribute throughout the entire extent of the DMN, some of them project mainly to either medial or lateral parts of the nucleus, thus suggesting that the organization of the DMN is comparable, at least in part, to that of the reticular formation of the pons and medulla, a region in which hodological differences between medial and lateral subdivisions are known to exist.

The superior and inferior salivatory nuclei in the rat.

The locations of the inferior and superior salivatory nuclei in the rat were determined by bathing the tympanic branch of the glossopharyngeal nerve and the chorda tympani with horseradish peroxidase (HRP) in different series of rats. The results demonstrated that the salivatory nuclei extend from the rostral medulla through the caudal pons. The inferior salivatory nucleus is composed of small to medium-sized neurons located in the ventrolateral portion of the reticular formation. It extends from a level at the rostral border of the nucleus ambiguus to the caudal part of the genu of the facial nerve. The superior salivatory nucleus differs in that it is composed of small to medium-sized neurons located in the dorsolateral portion of the reticular formation. It extends from a level at the caudal border of the facial nucleus through the genu of the facial nerve.

Efferent projections of the deep mesencephalic nucleus (pars lateralis) in the rat.

The projections of the lateral part of the deep mesencephalic nucleus (DMN) were traced by autoradiography and retrograde horseradish peroxidase (HRP) techniques. At the level of the DMN, projections from its lateral part crossed the midline and terminated in the medial and lateral part of the contralateral DMN. Furthermore, two labeled tracts passed rostrally from the lateral part of the DMN. One tract coursed dorsolaterally from the lateral DMN to terminate in the ipsilateral lateral thalamic nucleus. The second tract coursed ventrally and rostrally over the substantia nigra toward the ipsilateral zona incerta. At the caudal part of the zona incerta these fibers divided into two bundles. One bundle coursed superiorly to terminate bilaterally in the mediodorsal nucleus of the thalamus. The second bundle of fibers passed anteriorly to enter the ipsilateral zona incerta. Some of these fibers terminated upon neurons of the zona incerta and the ventromedial part of the subthalamic nucleus. The remaining fibers within the zona incerta coursed anteriorly to enter the internal capsule. These fibers terminated in the entopeduncular nucleus and medial part of the globus pallidus. These findings indicate that the lateral part of the DMN is likely to be involved in the ascending activating system of the reticular formation by connections with thalamic nuclei. Furthermore, the lateral part of the DMN may play a part in suprasegmental motor control via connections with rostral brain stem motor centers.

Efferent projections of the deep mesencephalic nucleus (pars medialis) in the rat.

The projections of the medial part of the deep mesencephalic nucleus (DMN) were traced by autoradiography and retrograde horseradish peroxidase (HRP) techniques. No ascending projections were observed from the medial part of the DMN; however, two groups of descending fibers were observed. One group crossed the midline and coursed to the caudal part of the red nucleus. At this point, these fibers divided into two distinct bundles. One bundle of fibers passed caudally to terminate in the contralateral pontine reticular nucleus, superior olive-trapezoid body complex, gigantocellular nucleus, and upper cervical spinal cord. The other bundle entered the medial longitudinal fasciculus (MLF) and coursed through the pons and medulla without termination, to enter the cervical spinal cord where terminations were noted in the dorsal horn. The other group of fibers from the medial DMN descended through the ipsilateral pons and medulla, projecting to the pontine reticular nucleus, superior olive-trapezoid body complex, gigantocellular nucleus, and upper cervical spinal cord. These findings indicate that the medial part of the DMN is likely to be involved in complex sensorimotor events via reticulobulbar and reticulospinal connections.

Pallidothalamic projections in the rat.

The purpose of this investigation was to examine pallidofugal projections to the thalamus of the rat using a selective silver degeneration method and autoradiography. Degenerating pallidothalamic projections emerging from the anterior part of the pallidum formed two diffuse groups of fibers; one coursed ventromedially directly through the internal capsule without forming a lenticular fasciculus and terminated uniformly in the reticular, VA, VD, VM and the medial part of the VE. The otehr group coursed ventrolaterally in the internal capsule and, without forming an ansa lenticularis, entered the zona incerta, swept medially through and ventral to the medial lemniscus, and terminated in the medial part of the VE and throughout the VD.