Dexamethasone significantly attenuates sub-arachnoid hemorrhage-induced elevation in cerebrospinal fluid citrulline and leukocytes.

AIM: Cerebral vasospasm is a leading cause of death and disability following aneurysmal subarachnoid hemorrhage (SAH). Nitric oxide (NO) is a potent mediator of vasodilation, and citrulline is a known contributor to NO production. The leukocytosis inflammatory response can increase vasoconstrictive compounds that may also contribute to vasospasm. Dexamethasone is a glucocorticosteroid commonly administered after SAH, which may alter the production of leukocytes and citrulline. The goal of this project was to study the effects of dexamethasone on leukocytosis, citrulline, and angiographic vasospasm. METHODS: Experimental SAH was induced in 18 New Zealand white rabbits. Intravenous dexamethasone was administered to one group (N.=9) at 2 mg/kg/day. A placebo group (N.=9) was given a saline infusion with otherwise identical procedures. CSF citrulline, leukocytes, protein, and glucose, as well as plasma citrulline were measured at baseline and 3 days post-SAH in a blinded fashion. Basilar artery angiography was performed at baseline and repeated 3 days post-SAH. RESULTS: The change in CSF citrulline from day 0 to day 3 was significantly lower in the dexamethasone group compared to controls (P=0.002). The change in CSF white blood cells was also significantly lower (P=0.005). There was no significant change in plasma citrulline levels or angiographic vasospasm. CONCLUSION: Dexamethasone significantly decreases CSF citrulline and CSF leukocytosis after experimental SAH. It is possible this could lead to a relative vasoconstriction and vasodilation, respectively. These processes could cancel-out opposing effects of dexamethasone on cerebral vasospasm, partially contributing to the recognized, multifactorial, inconsistent effects of glucocorticoids on vasospasm.

Prolonged constriction of sciatic nerve affecting oxidative stressors & antioxidant enzymes in rat.

BACKGROUND & OBJECTIVE: This study was carried out to determine the effects of reactive oxygen species in the balance between the pro-oxidant and antioxidant levels in experimental peripheral constriction injury induced by silver wire looping of sciatic nerve of rats. METHODS: Rats were divided into experimental group 1 (silver wire ligated) and group 2 (control, sham operated). Functional and behavioural activities were assessed by a modified Basso Beattie Bresnahan (BBB) locomotory rating scale. Mechanical pain intensity was measured with Randall and Selitto apparatus. Foot positioning, toe spread, paw withdrawal threshold and paw withdrawal latency were carried out on days 1, 3, 7, 14, 21 and 28 in rats with chronic pain. Oxidative stress markers such as malondialdehyde (MDA) and advanced oxidation protein products (AOPP) were measured along with antioxidants such as glutathione peroxidase (GPx), superoxide dismutase (SOD), catalase (CAT) and reduced glutathione (GSH) on day 30 after constriction in sciatic nerve, spinal cord, dorsal root ganglion, dorsal root and ventral root. RESULTS: Significant (P<0.05) increase in MDA, AOPP, SOD and GPx and decrease in the GSH and catalase activities in sciatic nerve, spinal cord, dorsal root ganglion, dorsal root and ventral root were observed in experimental group rats compared to control group. There was no recovery in foot positioning and toe spread. Reduced paw withdrawal threshold and paw withdrawal latency was observed in ligated rats compared to control rats. INTERPRETATION & CONCLUSION: Foot positioning, toe spread, paw withdrawal threshold and paw withdrawal latency with no recovery until day 30 confirmed locomotory deficits, hyperalgesia and neuronal impairment. Oxidative stress evidenced by increased MDA, AOPP and decreased GSH and catalase support the generation of reactive oxygen species in constriction model. The present experimental model for chronic pain induced by silver wire spirally coiled around sciatic nerve may be useful for future studies.

Origins and terminations of bulbospinal axons that contain serotonin and either enkephalin or substance-P in the North American opossum.

We have shown previously that some enkephalin, substance-P, and serotoninergic neurons in the medullary raphe and adjacent reticular formation project to the spinal cord in the opossum. In the present study we have combined the retrograde transport of True Blue and immunofluorescence histochemistry to determine whether methionine enkephalin or substance-P containing bulbospinal neurons are serotoninergic. Furthermore, we have used the same immunofluorescence protocol to determine whether spinal axons contain the same substances. Neurons that immunostained for both enkephalin and serotonin were observed in many brainstem nuclei. However, those that projected to the spinal cord were limited to the nuclei raphe magnus and obscurus, and the ventral part of nucleus reticularis gigantocellularis, pars ventralis. Neurons that immunostained for both substance P and serotonin were fewer in number, but some of the ones in the above nuclei and within the nucleus raphe pallidus, projected to the spinal cord. Spinal axons exhibiting both enkephalin- and serotonin-like immunoreactivity were observed in the superficial laminae of the dorsal horn, lamina X, and the intermediolateral cell column, whereas those showing both substance-P and serotonin-like immunoreactivity were seen primarily in lamina X, the intermediolateral cell column, and the ventral horn. Some of the axons in the ventral horn were in close apposition to presumed motoneurons. Comparison of the above results with those obtained from previous studies of bulbospinal projections has allowed us to infer the origins of axons that innervate different spinal targets.

Localization of enkephalinergic neurons in the dorsolateral pontine tegmentum projecting to the spinal cord of the cat.

The dorsolateral pontine tegmentum of the cat is known to contain enkephalinergic neurons, with most of the enkephalin co-contained in the catecholaminergic neurons; however, enkephalinergic cells projecting to the spinal cord have not been identified. This study employs retrograde transport of horseradish peroxidase in combination with methionine-enkephalin or tyrosine hydroxylase immunocytochemistry to 1) determine the locations of pontospinal enkephalinergic neurons and 2) compare these with the locations of pontospinal catecholaminergic neurons. Pontospinal enkephalinergic neurons were observed in the nuclei locus coeruleus and subcoeruleus and the Kölliker-Fuse nucleus. A high concentration of these neurons was evident in the Kölliker-Fuse nucleus when compared to the nuclei locus coeruleus and subcoeruleus (P less than .01). Both the enkephalinergic and catecholaminergic neurons projecting to the spinal cord were located in the same general areas of the dorsolateral pontine tegmentum and there was no significant difference in the mean diameters of these two neuronal types (P greater than .05). Quantitative data concerning the pontospinal enkephalinergic neurons correlated well with previous data on pontospinal catecholaminergic neurons (Reddy et al., Brain Res. 491:144-149, '89). A majority of the descending neurons from the dorsolateral pontine tegmentum contain enkephalin (72-80%) and catecholamine (80-87%). The observations suggest that enkephalin is contained in many of the pontospinal catecholaminergic neurons.

Conformationally restricted analogues of 1N,12N-bisethylspermine: synthesis and growth inhibitory effects on human tumor cell lines.

Eight analogues of 1N,12N-bisethylspermine (BES) with restricted conformations were synthesized in the search for new spermine mimetics with cytotoxic activities. By replacing the central butane segment of BES with a 1,2-disubstituted cyclopropane ring, a pair of cis/trans-isomers was obtained that introduced a spatial constraint in the otherwise freely mobile butane chain. An analogous pair of isomers was obtained when the butane segment was replaced with a 1, 2-disubstituted cyclobutane ring or with a 2-butene residue. The six new BES analogues thus obtained (three pairs of cis/trans-isomers) were growth inhibitory at low-micromolar concentrations against four human tumor cell lines (A549, HT-29, U251MG, and DU145) but were less growth inhibitory against two other human tumor cell lines (PC-3 and MCF7). 1N,12N-Bisethylspermyne, where the central butane segment of BES was replaced by the rigid 2-butyne segment, was devoid of growth inhibitory activity against five of the six human cell lines studied (DU145 being the only exception), a clear indication of the importance of conformational mobility at the 4N, 9N-butane segment of BES for its biological activity. When the butane segment was replaced by a benzene-1,2-dimethyl residue, the resulting BES analogue was devoid of growth inhibitory activity despite its cisoid conformation. The cytotoxicity of the analogues does not seem to be directly related to their uptake by the cells or to their effects on cellular polyamine levels. BES analogues with restricted conformations but which contained the equivalent of a two-carbon unit, rather than the natural four-carbon unit, at the central segment, such as 1,2-diaminocyclopropyl or 1, 2-diaminocyclobutyl derivatives, were devoid of growth inhibitory effects at the concentrations studied. The development of conformationally restricted polyamine analogues appears to show promise in the further quest for polyamine-related therapeutic agents with specificity of action.

Conformationally restricted analogues of 1N,14N-bisethylhomospermine (BE-4-4-4): synthesis and growth inhibitory effects on human prostate cancer cells.

Twelve analogues of 1N,14N-bisethylhomospermine (BE-4-4-4) with restricted conformations were synthesized in the search for cancer chemotherapeutic agents with higher cytotoxic activities and lower systemic toxicities than BE-4-4-4. The central butane segment of BE-4-4-4 was replaced with a 1,2-substituted cyclopropane ring, a 1,2-substituted cyclobutane ring, and a 2-butene residue. In each case, the cis/trans-isomeric pair was synthesized. Cis-monounsaturation(s) was also introduced at the outer butane segment(s) of BE-4-4-4. The two possible cis-dienes and a cis-triene formally derived from the tetraazaeicosane skeleton of BE-4-4-4 were also prepared. Four cultured human prostate cancer cell lines (LnCap, DU145, DuPro, and PC-3) were treated with the new tetramines to examine their effects on cell growth with a MTT assay. One representative cell line (DuPro) was selected to further study the cellular uptake of the novel tetramines, their effects on intracellular polyamine pools, and their cytotoxicity. All tetramines entered the cells, reduced cellular putrescine and spermidine pools while exerting only a small effect on the spermine pool, inhibited cell growth, and killed 2-3 logs of cells after 6 days of treatment at 10 microM. Four new tetramines, the two cyclopropyl isomers, the trans-cyclobutyl isomer, and the (5Z)-tetraazaeicosene, were more cytotoxic than their saturated counterpart (BE-4-4-4). Their cytotoxicity, however, could not be correlated either with their cellular uptake or with their ability to deplete intracellular polyamine pools. We attribute their cytotoxicity to their specific molecular structures. The cytotoxicity was markedly reduced when the central butane segment was deprived of its rotational freedom by replacing it with a double bond. Introduction of a triple bond or a benzene-1,2-dimethyl residue at the central segment of the polyamine chain, led to complete loss of biological activity. The conformationally restricted alicyclic derivatives were not only more cytotoxic than was the freely rotating BE-4-4-4 by several orders of magnitude but also had much lower systemic toxicities than the latter. Thus, we obtained new tetramines with a wider therapeutic window than BE-4-4-4.

cis-Unsaturated analogues of 3,8,13,18,23-pentaazapentacosane (BE-4-4-4-4): synthesis and growth inhibitory effects on human prostate cancer cell lines.

From the results of our previous physicochemical studies of polyamine-nucleic acid interactions, we concluded that polyamine analogues in cisoidal conformation are capable of wrapping around the major groove of the double helix, of displacing natural polyamines from their nucleic acid binding sites, and of inhibiting cell division. On the basis of this hypothesis, nine unsaturated pentamines, formally derived from the cytotoxic pentamine 3,8,13,18,23-pentaazapentacosane (BE-4-4-4-4), were prepared in an attempt to increase antineoplastic activity. Cis-double bonds were introduced in all possible sites in the saturated pentaazapentacosane structure of BE-4-4-4-4 to yield two pentacosenes, four pentacosadienes, two pentacosatrienes, and one pentacosatetraene. Cis-double bonds should also provide good targets for mixed-function oxidases that might eliminate the accumulation of unsaturated pentamines in serum, thereby reducing systemic toxicity in animals. We determined the ability of these new pentamines to inhibit growth in four cultured human prostate cancer cell lines (LnCap, DU145, PC-3, and DuPro) using a MTT assay. LnCap and DU145 cells were very sensitive, PC-3 cells were relatively resistant, and DuPro cells were intermediate in sensitivity to most of these synthetic pentamines. In all cell lines, pentamines that had unsaturation(s) at the end of the chain showed the highest cell growth inhibitory effects. The cellular uptake, effects on cellular polyamine levels, and cytotoxicity of these pentamines on one representative prostate cancer cell line (DuPro) were further examined with a colony-forming efficiency (CFE) assay. The pentamines with unsaturation(s) at the end of the chain were once again the most cytotoxic among both the saturated (BE-4-4-4-4) and unsaturated analogues. Appreciable amounts of all pentamines entered DuPro cells and depleted cellular polyamine pools by day 6 of treatment. For most pentamines, however, cell growth inhibitory and cytotoxic effects could not be directly correlated either with their cellular uptake or with their ability to deplete cellular polyamine pools. The position of the double bonds in the aliphatic backbone seems to be the most important determinant of cytotoxicity. For some pentamines, however, depletion of cellular polyamines may add to their efficacy.

Localization of glutamatergic neurons in the dorsolateral pontine tegmentum projecting to the spinal cord of the cat with a proposed role of glutamate on lumbar motoneuron activity.

Glutamate is considered to be a major excitatory neurotransmitter in the central nervous system. The presence of glutamate-like immunoreactive neurons in the rodent locus coeruleus has been reported previously. In this study we used both immunohistochemical and electrophysiological techniques to answer two major questions: (1) Is there any glutamate-like immunoreactivity in the catecholaminergic coeruleospinal system of the cat? (2) What is the physiological role, if any, of glutamate in descending locus coeruleus control of spinal motoneurons? Following injections of rhodamine-labeled latex microspheres or Fast Blue into the seventh lumbar segment of the spinal cord of the cat, retrogradely labeled cells were found throughout the rostrocaudal extent of the dorsolateral pontine tegmentum. They were primarily observed in the nucleus locus coeruleus and the Kolliker-Fuse nucleus. Some labeled cells were also present in the nucleus subcoeruleus and, to a lesser extent, in the parabrachial nuclei. Data from immunohistochemical studies indicate that 86% of all dorsolateral pontine tegmentum neurons that project to the spinal cord contain glutamate-like immunoreactivity, and 77% co-contain both glutamate- and tyrosine hydroxylase-like immunoreactivity. Electrical stimulation (four pulses of 500 microseconds duration at 500 Hz; intensity = 50-200 microA) of the locus coeruleus, in decerebrate cats, consistently induced lumbar motoneuron discharges recordable ipsilaterally as ventral root responses. These motoneuronal responses were reversibly antagonized following chemical inactivation of noradrenergic locus coeruleus neurons by local infusion of the alpha 2-adrenergic agonist clonidine, suggesting the locus coeruleus neurons to be the main source of evoked ventral root responses. Additionally, the evoked ventral root responses were reversibly reduced by 34.20 +/- 4.45% (mean +/- S.E.M.) upon intraspinal injections of the non-N-methyl-D-aspartate receptor antagonist, 6-cyano-7-nitroquinoxaline-2,3-dione, into the ventral horn of seventh lumbar spinal cord segment (three to four injections, 20 nmol in 0.2 microliter of 0.1 M Tris-buffered saline for each injection). Similar volumes of vehicle injections had no significant effect on the locus coeruleus-evoked ventral root responses. These ventral root responses were also partially blocked (62.30 +/- 11.76%) by intravenous administration of the alpha 1-adrenergic receptor antagonist prazosin (20 micrograms/kg). In the light of several anatomical reports of noradrenergic and glutamatergic terminals in close contact with spinal motoneurons, our present findings suggest that the locus coeruleus-evoked ventral root response probably involves the synaptic release of both norepinephrine and glutamate onto lumbar motoneurons.(ABSTRACT TRUNCATED AT 400 WORDS)

Pontospinal transmitters and their distribution.

The dorsolateral pontine tegmentum of the cat is known to contain a large population of catecholaminergic neurons. Additionally, several studies have also shown the presence of other neurochemicals (acetylcholine, enkephalin, neuropeptide Y, serotonin, somatostatin and substance P). In this study, we have employed retrograde transport of horseradish peroxidase in combination with immunocytochemistry to determine the locations of pontospinal neurons which contain catecholamine, enkephalin, neuropeptide Y, and serotonin. Furthermore, we have combined the retrograde transport of Fast Blue and immunofluorescence histochemistry to determine whether enkephalin-containing neurons are catecholaminergic. All pontospinal neurons, irrespective of the neurochemical content, were observed in the ventral and lateral parts of the dorsolateral pontine tegmentum at coronal levels P1.8-P4.0. These neurons were located in the nuclei locus coeruleus alpha and subcoeruleus and the Kölliker-Fuse nucleus. A high concentration of these neurons was evident in the Kölliker-Fuse nucleus when compared to the nuclei locus coeruleus alpha and subcoeruleus. Quantitative data have revealed that enkephalin is contained in a large proportion of the pontospinal catecholaminergic neurons (75%). The observations suggest that catecholaminergic neurons may contain one or more putative peptide neurotransmitters.

Immunohistochemical evidence for coexistence of methionine-enkephalin and tyrosine hydroxylase in neurons of the locus coeruleus complex projecting to the spinal cord of the cat.

Previous studies have revealed the presence of pontospinal neurons with either methionine-enkephalin- or tyrosine hydroxylase-like immunoreactivity in the dorsolateral pontine tegmentum of the cat. Using a combined fast blue retrograde transport technique and simultaneous immunofluorescence histochemistry, the present study was designed to reveal the coexistence of enkephalin and tyrosine hydroxylase in cat coerulospinal neurons and to determine if and to what extent the coerulospinal pathway is heterogeneous. Fast blue-labelled neurons with tyrosine hydroxylase- and enkephalin-like immunoreactivities were found in the nucleus locus coeruleus, nucleus subcoeruleus, Kölliker-Fuse nucleus, and the medial and lateral parabrachial nuclei. Approximately 87% of tyrosine hydroxylase-like immunoreactive neurons had enkephalin-like immunoreactivity, whereas about 76% of the enkephalin-like immunoreactive neurons had tyrosine hydroxylase-like immunoreactivity. About 71% of all coerulospinal neurons exhibited both tyrosine hydroxylase- and enkephalin-like immunoreactivities. These findings indicate that coerulospinal activity may lead to spinal cord effects reflecting both norepinephrine and enkephalin activity in most cases but do not rule out each transmitter's isolated functions.

Anatomical evidence for the presence of glutamate or enkephalin in noradrenergic projection neurons of the locus coeruleus.

This paper reviews the anatomical evidence for the presence of glutamate (GLU) in noradrenergic neurons of the nucleus locus coeruleus (LC) and adjacent nuclei in the dorsolateral pontine tegmentum (DLPT) that project to the spinal cord, cerebellum, or cerebral cortex. Additionally, the evidence for the existence of methionine-enkephalin (ENK) in noradrenergic neurons of the DLPT that project to the spinal cord of the cat is reviewed. In these studies, we have combined the retrograde transport of either Fast Blue (FB), rhodamine labeled latex microspheres (MS), or rhodamine labeled dextran and indirect immunofluorescence histochemistry to determine whether the neurons that contain tyrosine hydroxylase (TH) and project to these terminal fields also contain GLU or ENK. The neurons of the cat that project to the spinal cord, cerebellum, and neocortex were observed in the nucleus LC and Kölliker-Fuse (KF) nucleus. They were also present, to a lesser extent, in the nucleus subcoeruleus (SC) and nuclei parabrachialis medialis (PBM) and lateralis (PBL). In the rat the majority of the neurons that project to the neocortex and hippocampus were located in the nucleus LC. Our data revealed a major proportion of these neurons to be immunostained for both GLU and TH (cat, rat), or ENK and TH (cat). Functional implications of such colocalized neurochemicals within individual LC projection neurons are discussed.

Coerulospinal cells containing neuropeptide Y in the cat.

Spinally projecting neuropeptide Y (NPY)-immunoreactive cells were sought in the feline locus coeruleus (LC) nuclear complex after horseradish peroxidase (HRP) injection into the lumbar cord; HRP injection was followed by intracerebroventricular colchicine administration. Our results revealed that a significant number (approximately 20% of all descending cells from the LC complex) of spinally projecting NPY-immunoreactive neurons arise from the LC alpha, the subcoeruleus and the Kölliker-Fuse nuclei. Other nonspinally projecting NPY-containing cells were also evident in the laterodorsal tegmental nucleus and the LCd, in addition to those occurring in the aforementioned LC nuclear complex.

Spinally projecting noradrenergic neurons of the dorsolateral pontine tegmentum: a combined immunocytochemical and retrograde labeling study.

Two to three days following injection of horseradish peroxidase (HRP) into the spinal cords of 5 cats, the animals were sacrificed and perfused, and the brainstems removed and sectioned. The sections were then processed for HRP and, immunocytochemically, for tyrosine hydroxylase (TH). The dorsolateral pontine tegmentum was divided into the locus coeruleus, subcoeruleus and Kölliker-Fuse nucleus; the mean percentage of pontospinal neurons containing TH were found to be 85.5 +/- 2.5 (S.E.M.), 79.6 +/- 5.6 and 87.1 +/- 3.1, respectively. The cell diameters of locus coeruleus cells were also measured.

Differential labeling of the vestibular complex following unilateral injections of horseradish peroxidase into the cat and rat locus coeruleus.

Following unilateral injections of horseradish peroxidase into the cat and rat locus coeruleus (LC), bilateral retrograde labeling was evident in all 4 vestibular nuclei. In both species, the major contributions of LC afferents arose from the ipsilateral rather than the contralateral vestibular nuclei. Quantitative analysis in the rat has indicated that approximately 90% of the total vestibular-LC projections originated from the ipsilateral vestibular nuclear complex, with 10% from the contralateral. The frequency distribution was in the order of lateral greater than medial greater than superior greater than inferior vestibular nuclei.

Effect of chronic lithium chloride on membrane adenosine triphosphatases in certain postural muscles of rats.

Lithium has been extensively used as an antidepressant in the treatment of manic depressive disorders requiring chronic administration. Here, we report a study of the effect of long-term lithium treatment on the activities of membrane adenosine triphosphatases (ATPases) in certain postural muscles of rat. Specifically, Ca(2+)-ATPase, Na+,K(+)-ATPase and Mg(2+)-ATPase activities were measured in the soleus, extensor digitorum longus and plantaris muscles following 6 weeks of treatment with LiCl. Increases were observed in the Na+,K(+)-ATPase activity whereas the Mg(2+)-ATPase activity decreased with prolonged LiCl treatment. The most pronounced effect was a highly significant (P < 0.001) increase in the mitochondrial Ca(2+)-ATPase and Na+,K(+)-ATPase activity to almost 50-100% above the control. The increases in the mitochondrial Ca(2+)-ATPase activity of extensor digitorum longus and plantaris were 70% and 100%, respectively. The corresponding increases in the Na+,K(+)-ATPase activity were 127%, 99% and 87% for soleus, extensor digitorum longus and plantaris, respectively. Irrespective of the differences in the fiber pattern and physiological function, all three muscles responded in a similar way to Li+. The changes in the membrane ATPases reflect a deranged ATP turnover, thus affecting the overall energy state of the animal. Based on these results, we hypothesize that Li+ produces its effects by interfering with cation transport processes. Since Li+ affects the neural excitability of the cell it is suggested that the stimulation of the ATPases may be important in the psychotropic properties of the ion.

Serotonergic and non-serotonergic raphe neurons projecting to the feline lumbar and cervical spinal cord: a quantitative horseradish peroxidase-immunocytochemical study.

A quantitative study of raphe-spinal neurons and serotonergic neurons in 3 medullary raphe nuclei in the cat indicates that more than 80% of the raphe-spinal neurons that project to the spinal cord are serotonergic raphe-spinal neurons in each of the nuclei. More than 85% of the descending raphe-spinal neurons in the two caudal nuclei, nucleus raphe pallidus and nucleus raphe obscurus, are serotonergic, whereas 75% of the raphe-spinal neurons in the more rostrally placed nucleus raphe magnus contain serotonin (5-HT). These results are discussed in relation to descending systems containing both neuropeptides and 5-HT and collateralizing to several spinal segments.

Immunohistochemical localization of glutamate-containing neurons in the lateral reticular nucleus projecting to the cerebellar vermis in the kitten.

The lateral reticular nucleus (LRN) and afferents to the cerebellum are known to contain glutamate-like immunoreactive (Glu-LI) neurons and axons, respectively. However, such a direct link between the Glu-LI LRN neurons and the cerebellar vermal cortex has not been identified. In this study we have combined the retrograde transport of rhodamine labeled latex microspheres and immunofluorescence histochemistry to determine the locations of Glu-LI neurons of the kitten reticulocerebellar system. Following microsphere injections into the cerebellar vermis (lobules V-VII), retrogradely labeled neurons were encountered throughout the rostrocaudal extent of the LRN. More than 60% (n = 3 kittens) of these retrogradely labeled neurons were immunostained for Glu-like immunoreactivity. Our observations of the Glu-like immunoreactivity in a majority of the reticulocerebellar neurons suggest that Glu in these neurons may participate in LRN's control of target neuron activities in the cerebellar vermis of kittens.

Hippocampal beta-amyloid reduces locus coeruleus glutamate and tyrosine hydroxylase.

The effects of intrahippocampally injected beta-amyloid protein (beta-AP) on glutamate- (Glu) and tyrosine hydroxylase (TH)-like immunoreactivities in the neurons of the locus coeruleus (LC) were studied in rats. A synthetic peptide or the vehicle alone was injected into the hippocampus as controls. All injections were made once a week (two or three injections; 3 nmol in 2 microliters of distilled water). Fluorescent microspheres (either alone or with one of the peptides) were also injected into the hippocampus to identify coeruleo-hippocampal neurons. The results revealed cell loss in the hippocampus at the site near beta-AP or control peptide deposition. Furthermore, in beta-AP/microsphere injected animals, only 22.4% and 49.6% of hippocampal projection neurons contained Glu and TH, respectively, compared to 88.4% and 85.3% in the animals that received control peptide with microspheres. Our results suggest that beta-AP has an effect on noradrenergic cells whose axons project to the hippocampus. These effects may contribute to the TH cell loss in the LC of Alzheimer's brains.

Existence of glutamate in noradrenergic locus coeruleus neurons of rodents.

This study distinguished three types of immunolabeled neurons in nucleus locus coeruleus (LC) of the rat and mouse: cells single labeled either for tyrosine hydroxylase-like immunoreactivity (TH-LI) or glutamate (Glu)-LI, and those double labeled for both antigens. Although the double labeled neurons tend to be located in the middle and ventral thirds of the rat LC nucleus, throughout its rostrocaudal extent, such feature was not apparent in the mouse. Quantitatively a majority of neurons cocontaining TH- and Glu-LI were commonly observed in the rat (62%) and mouse (77%) LC. Additional studies utilizing the combined retrograde and immunohistochemical labeling revealed that such a high incidence of coexistence of the TH- and Glu-LI was also represented by coeruleocortical neurons in the rat (69% and 75% of all ipsilateral and contralateral projection cells, respectively). A possible role of coeruleocortical neurons involvement in Glu- and norepinephrine-mediated target neuron dysfunction is discussed.

Cotransmitter-mediated locus coeruleus action on motoneurons.

This article reviews evidence for a direct noradrenergic projection from the dorsolateral pontine tegmentum (DLPT) to spinal motoneurons. The existence of this direct pathway was first inferred by the observation that antidromically evoked responses occur in single cells in the locus coeruleus (LC), a region within the DLPT, following electrical stimulation of the ventral horn of the lumbar spinal cord of the cat. We subsequently confirmed that there is a direct noradrenergic pathway from the LC and adjacent regions of the DLPT to the lumbar ventral horn using anatomical studies that combined retrograde tracing with immunohistochemical identification of neurotransmitters. These anatomical studies further revealed that many of the noradrenergic neurons in the LC and adjacent regions of the DLPT of the cat that send projections to the spinal cord ventral horn also contain colocalized glutamate (Glu) or enkephalin (ENK). Recent studies from our laboratory suggest that Glu and ENK may function as cotransmitters with norepinephrine (NE) in the descending pathway from the DLPT. Electrical stimulation of the LC evokes a depolarizing response in spinal motoneurons that is only partially blocked by alpha 1 adrenergic antagonists. In addition, NE mimicks only the slowly developing and not the fast component of LC-evoked depolarization. Furthermore, the depolarization evoked by LC stimulation is accompanied by a decrease in membrane resistance, whereas that evoked by NE is accompanied by an increased resistance. That Glu may be a second neurotransmitter involved in LC excitation of motoneurons is supported by our observation that the excitatory response evoked in spinal cord ventral roots by electrical stimulation of the LC is attenuated by a non-N-methyl-D-aspartate glutamatergic antagonist. ENK may participate as a cotransmitter with NE to mediate LC effects on lumbar monosynaptic reflex (MSR) amplitude. Electrical stimulation of the LC has a biphasic effect on MSR amplitude, facilitation followed by inhibition. Adrenergic antagonists block only the facilitator effect of LC stimulation on MSR amplitude, whereas the ENK antagonist naloxone reverses the inhibition. The chemical heterogeneity of the cat DLPT system and the differential responses of motoneurons to the individual cotransmitters help to explain the diversity of postsynaptic potentials that occur following LC stimuli.