Recent Publications by CFE Educators

In this report, we describe an HPLC with electrochemical detection assay for the simultaneous measurement of levels of morphine, serotonin, 5-hydroxyindole-3-acetic acid, and homovanillic acid in dialysates of various brain areas and CSF in the awake rat. Morphine could be detected in the dialysates after a single intraperitoneal injection, with doses as low as 1.0 mg/kg. The time course of extracellular morphine content in the lateral hypothalamus, striatum, cerebellum, periaqueductal gray, and dorsal horn of the spinal cord and in CSF, from the ventricles and cisterna magna, was similar. We detected morphine in the first 15-min sample, and levels peaked 45-60 min after injection. Maximal dialysate levels, however, varied with the type of dialysis probe used and the area sampled. The most efficient in vivo recovery was in CSF dialysates from the cisterna magna, presumably because of minimal tissue interference with the dialysis probe. For this reason, the cisterna is an ideal region for sampling CSF. Morphine had no significant effect on the extracellular concentrations of serotonin in any of the areas studied and did not modify or only slightly increased levels of tissue metabolites; however, morphine markedly increased the CSF levels of 5-hydroxyindole-3-acetic acid and homovanillic acid. Because microdialysis in freely moving animals permits assessment of the behavioral effects of morphine while continuously monitoring the drug levels in discrete brain regions, this approach will greatly facilitate future studies of the neurochemical basis of morphine's effects in the brain.

Bombesin-like peptides (BLP) produced by pulmonary neuroendocrine cells have many physiological actions which are relevant to the pathobiology of cigarette smoking. The objectives of this study were to determine whether cigarette smokers excrete increased levels of BLP in their urine compared with nonsmokers, to determine the relationship between BLP levels in urine and bronchoalveolar lavage (BAL) fluid, and whether urinary BLP levels are merely a reflection of exposure to cigarette smoke. Simultaneous BAL fluid and urine samples were obtained from ten clinically normal smokers and 22 normal nonsmoker volunteers. Urine samples were also obtained from 39 normal smokers and 30 normal nonsmokers who did not have BAL performed. BLP levels were measured in urine and BAL fluid using an enzyme-linked immunoassay. Expired air content of carbon monoxide, which reflects recent exposure to cigarette smoke, was determined in 34 of the clinically normal smokers and correlated with urinary BLP levels. We found that, in addition to having increased BLP levels in BAL fluid (P = 0.04), asymptomatic cigarette smokers also have increased BLP levels in their urine compared with normal nonsmokers (P = 0.007). Of note, a subgroup of smokers have markedly increased BLP levels which do not overlap with the nonsmokers. Urinary BLP levels correlated with expired air content of carbon monoxide (r = 0.49, P less than 0.01). However, not all smokers with increased expired air content of carbon monoxide exhibited increased BLP levels. Finally, all smokers with detectable BLP levels in BAL fluid had detectable urinary BLP levels, and there was a positive correlation between BLP levels in urine and BAL fluid (r = 0.625, P less than 0.001). We conclude that a subgroup of asymptomatic cigarette smokers exhibited increased BLP levels, measurable in both urine and BAL fluid, which precede the onset of clinically detectable disease and which are not strictly dependent on smoking intensity. We speculate that smokers with increased BLP levels may have a greater risk for smoking-related diseases.

Pulmonary histiocytosis X is an uncommon but important cause of pulmonary fibrosis and honeycombing in young adults. This article reviews the pathologic, clinical, radiographic, and high-resolution computed tomographic (HRCT) features of pulmonary histiocytosis X, focusing on differential diagnosis and disease progression. The main pathologic feature of pulmonary histiocytosis X is peribronchiolar inflammation, leading to fibrosis and cyst formation. Although the diagnosis of pulmonary histiocytosis X may be suspected on the basis of chest radiographic findings, the HRCT findings of predominantly upper lobe nodules and cysts are virtually pathognomonic of this disorder. As pulmonary histiocytosis X progresses, the nodules decrease in number, leaving multiple thin-walled cysts. HRCT can be useful in visually estimating the proportion of lung involved and is also valuable in distinguishing histiocytosis from other disorders that produce nodules or cysts. Chest HRCT helps confirm the pattern seen at radiography and is valuable in establishing the correct diagnosis.

The synaptic relationships between gamma-aminobutyric acid (GABA)-immunoreactive and enkephalin-immunoreactive profiles in the cat spinal cord were examined using combined pre-embedding immunoperoxidase and post-embedding immunogold electron microscopic immunocytochemistry. Although colchicine was not used, enkephalin-immunoreactive somata and dendrites were detected in regions associated with nociceptive transmission, including laminae I, II, V and X. In each of these laminae, many GABA-immunoreactive terminals were found presynaptic to enkephalin-immunoreactive cell bodies and dendrites. We propose that disinhibition of opioid-containing neurons may be a common feature of pain-related circuits in the cat spinal cord.

This study investigated the regulation of serotonin (5-HT) and its major metabolite 5-hydroxyindoleacetic acid (5-HIAA) in the dorsal spinal cord of awake, freely moving rats, using microdialysis coupled to HPLC with electrochemical detection and tested the hypothesis that opioids exert their analgesic effect in part through the increased release of 5-HT in the dorsal horn. A dialysis tube was placed transversely at the L4 segment of the dorsal spinal cord and the basal concentration of 5-HT in the dialysate was characterized by infusion of a variety of substances through the dialysis probe: tetrodotoxin (TTX), KCl, imipramine, fluoxetine and amphetamine (AMPH). To evaluate the contribution of opioids, we also studied the effects of either systemic or intracerebroventricular (i.c.v.) injection of morphine or DAMGO. Extracellular concentrations of 5-HT and 5-HIAA were partially and reversibly reduced by TTX. In the presence of KCl, imipramine, fluoxetine or AMPH, 5-HT levels significantly increased. Under these conditions, extracellular 5-HIAA levels usually decreased. By contrast, the effects of opioids on 5-HT concentrations were highly variable. Low doses of morphine administered systemically increased 5-HT concentrations in only 3 of 6 rats. This was paralleled by a decrease in 5-HIAA. Higher doses of morphine, alone or in the presence of fluoxetine, did not change 5-HT concentrations. Intracerebroventricular injection of morphine or DAMGO increased the extracellular concentrations of 5-HT in only about one third of the animals. After intracerebroventricular opioid injection, extracellular concentrations of 5-HIAA either decreased by about 20% or did not change.(ABSTRACT TRUNCATED AT 250 WORDS)

Immunohistochemical visualization of Fos protein, the nuclear phosphoprotein product of the early-immediate gene c-fos, permits identification of populations of neurons that are activated in response to a variety of stimuli. This study examined the distribution of Fos-like immunoreactive (FLI) neurons in the spinal cord and the nucleus tractus solitarii (NTS) of the caudal medulla evoked by a noxious visceral stimulus in the unanesthetized rat. It also compared the inhibition of pain behavior and Fos expression by a mu-selective opioid agonist, morphine, and a kappa-selective opioid agonist, U-50,488. Intraperitoneal injection of 3.5% acetic acid in the unanesthetized rat evoked the expression of FLI in a discrete population of spinal cord neurons, the distribution of which closely mirrored the spinal terminations of visceral primary afferents. Specifically, FLI neurons were concentrated in laminae I, IIo, V, VII, and X. Large numbers of Fos-immunoreactive neurons were also present in the NTS of the caudal medulla, most likely as a result of spinosolitary tract and vaginal afferent input. The number of labeled neurons in both the spinal cord and the NTS was significantly correlated with the number of abdominal stretches, a pain behavior measure. Both morphine (1-10 mg/kg s.c.) and U-50,488 (3-30 mg/kg s.c.) produced a dose-dependent inhibition of the pain behavior in these animals and a dose-dependent suppression of the number of FLI neurons in both the spinal cord and in the NTS; complete suppression of FLI neurons was, however, not necessary for the production of antinociception. Furthermore, although equianalgesic doses of morphine and U-50,488 reduced the number of labelled neurons in the spinal cord to a comparable extent, morphine reduced the number of immunoreactive neurons in the NTS to a greater extent than did U-50,488. These results suggest that morphine and U-50,488 have comparable effects on the transmission of visceral nociceptive messages by spinal neurons, but differentially affect the autonomic response to noxious visceral stimuli.

Although it has been proposed that the locus coeruleus is the predominant, if not exclusive, brainstem origin of the noradrenergic innervation of the spinal dorsal horn, pharmacological studies argue otherwise. In this study we made localized injections of the retrograde tracer wheatgerm agglutinin conjugated to apo-horseradish peroxidase gold (WGA:apoHRP-Au), in conjunction with immunocytochemical labeling for tyrosine hydroxylase (TH) or serotonin (5-HT), to identify the brainstem source of the noradrenaline (NA) and 5-HT innervation of the dorsal horn of the rat. Our studies were concentrated in the C5 spinal segment. The pattern of labeling was only studied in animals in which the tracer injection was restricted to the dorsal horn. In these rats, TH-immunoreactive neurons in widespread regions of the brainstem, including the locus coeruleus, subcoeruleus, A5, and A7 cell groups, were found to project to the dorsal horn. In terms of absolute numbers of double-labeled cells, no one noradrenergic cell group predominated. As expected, dorsal-horn-projecting 5-HT-immunoreactive neurons were found within the 5-HT populations of the rostroventromedial medulla and caudal pons, including the nucleus raphe magnus, nucleus paragigantocellularis (PGi), and ventral portions of the nucleus gigantocellularis (Gi). The majority of retrogradely labeled 5-HT-immunoreactive cells were, however, located off the midline, in the ipsilateral PGi and ventral Gi. Finally, a large number of retrogradely labeled, non-5-HT cells were found intermingled among the 5-HT cells of this region. Our results provide evidence that the noradrenergic regulation of nociceptive transmission at the spinal cord level arises from direct spinal projections of several brainstem noradrenergic cell groups.

Recently, Bennett and Xie reported that when the sciatic nerve of the rat is ligated loosely, the rat develops a pain syndrome with many features similar to those observed in neuropathic pain states in man. Anatomical and physiological studies to date indicate that the major pathology is a loss of large diameter myelinated fibers distal to the ligatures, with more subtle changes in small myelinated fibers. With a view to evaluating possible changes in the unmyelinated fibers, we have performed an electron microscopic analysis of the sciatic nerve 2 weeks after four ligatures were applied, at which time the animals displayed profound hyperalgesia and mechanical and thermal allodynia. Cross-sectional photomontages of regions proximal and distal to the ligatures were studied. Consistent with light microscopic and electrophysiological studies, we found a near complete loss of large myelinated fibers distal to the ligatures. Phagocytosis of large fibers was common. There was also considerable variation in the damage to small myelinated fibers. In some fascicles many small (less than 3 microns) myelinated axons remained; in other fascicles none could be detected. Importantly, we also found significant changes in the unmyelinated fiber spectrum. Counts of unmyelinated axons revealed a 34% and 71% decrease in the distal compared to the proximal nerve, in the two rats studied. The large clusters of unmyelinated axons that characterize normal nerve (and the nerve proximal to the ligatures) were rarely found distally. Rather, many of the unmyelinated axons coursed singly or in very loose bundles. Many of the surviving axons were shrunken and distorted, although still in contact with Schwann cells.(ABSTRACT TRUNCATED AT 250 WORDS)