beta-Endorphin-induced stimulation of central sympathetic outflow: beta-endorphin increases plasma concentrations of epinephrine, norepinephrine, and dopamine in rats.

Intracisternal administration of synthetic human beta-endorphin (0.058-7.25 nmol) in chronically cannulated, conscious, freely moving, adult male rats increased plasma concentrations of epinephrine, norepinephrine, and dopamine in a dose-related manner. Epinephrine secretion was the most sensitive to the stimulatory effect of intracerebral beta-endorphin; plasma epinephrine increased transiently in response to 0.058 nmol. Of the three catecholamines, plasma epinephrine showed the greatest and most rapid response to the largest dose (7.25 nmol) studied. Plasma norepinephrine increased significantly in response to 1.45 nmol, peaking later than plasma epinephrine. Plasma dopamine increased only in response to the highest dose examined. These beta-endorphin effects on plasma catecholamines were inhibited by intraarterial naloxone (1.1 mumol/kg), supporting mediation at opioid receptors. Pretreatment with the ganglionic blocking agent chlorisondamine inhibited the responses of all three catecholamines to intracisternal beta-endorphin. Bilateral adrenal denervation completely prevented the plasma epinephrine response to beta-endorphin and blunted the plasma norepinephrine and dopamine responses. Prior intracisternal administration of hemicholinium-3 blocked the plasma responses of all three catecholamines to intracisternal beta-endorphin, providing evidence for the involvement of central cholinergic neurons in the mechanism mediating beta-endorphin-induced increases in plasma catecholamines. The data are consistent with the hypothesis that endorphins act at a presently unknown brain site(s) to increase the central sympathetic outflow to adrenal medulla and peripheral sympathetic nerve endings, thus stimulating peripheral catecholamine release and increasing plasma concentrations of epinephrine, norepinephrine, and dopamine.

Role for brain corticotropin-releasing factor in the weight-reducing effects of chronic fenfluramine treatment in rats.

Fenfluramine is an amphetamine derivative which is used as a weight-reducing agent in the treatment of obesity. It has been postulated that fenfluramine affects brain serotonin (5HT) neurons resulting in decreased food intake and altered autonomic outflow which, in turn, increases metabolism. CRF decreases food intake and, in addition, has been demonstrated to reduce body weight in genetically obese rats through selective activation of sympathetic and inhibition of parasympathetic outflows. Because 5HT is a potent CRF secretagogue, we tested the hypothesis that the weight-reducing effects of fenfluramine administration may be mediated, in part, through altered CRF secretion. Chronic fenfluramine treatment (1-24 mg/kg sc, twice daily, 4 days) resulted in a dose-dependent decrease in hypothalamic CRF concentration at 30 min after the final drug injection and was accompanied by a significant reciprocal increase in plasma corticosterone concentration. These data suggest that the decrease in hypothalamic CRF was a consequence of increased CRF secretion. These changes in hypothalamic CRF and plasma corticosterone correlated with brain fenfluramine levels. In contrast, high dose fenfluramine treatment significantly increased hippocampus, midbrain, and spinal cord CRF concentrations whereas levels in cerebral cortex, caudate putamen, thalamus, pons/medulla, and cerebellum were unaffected. There was no effect of this fenfluramine treatment protocol on regional brain TRH or neurotensin concentrations. In keeping with the well known development of tolerance to the weight-reducing effects of fenfluramine, chronic fenfluramine treatment resulted in lesser increases in corticosterone secretion than after acute treatment. Whereas weight loss observed after chronic fenfluramine treatment was associated with stimulation of hypothalamic-pituitary-adrenocortical hormone secretion, the weight-recovery phase after cessation of drug treatment was associated with decreased levels of plasma corticosterone. These data, demonstrating fenfluramine-induced alterations in brain CRF and plasma corticosterone, suggest that CRF may represent an important endogenous transmitter which mediates the weight-reducing effects of the drug.

Muscarinic acetylcholine receptor subtype mRNA expression and ligand binding in the aged rat forebrain.

Previous studies indicate that a 20-30% decline in muscarinic acetylcholine receptor binding occurs in localized areas of rat brain during aging. In this study, reduced [3H]-quinuclidinyl benzilate binding was observed in striata from 24-25-month-old rats relative to 5-6-month-old animals using homogenate binding assays. To determine if the decline in receptor concentration occurs as a result of decreased receptor synthesis, the expression of the m1, m3, and m4 muscarinic receptor mRNAs as well as [3H]-QNB binding were determined in adjacent sections of young and old male rats using in situ hybridization and in vitro receptor autoradiography respectively. A significant decline in collective muscarinic receptor binding as assessed by [3H]-QNB was observed in the caudate putamen, olfactory tubercle, nucleus accumbens, and several frontal and parietal cortical areas. The only difference observed in muscarinic mRNA expression for any of the three subtypes examined was a decline in m1 hybridization in the olfactory tubercle. The results of this study demonstrate that the regional brain areas displaying age-related decreases in receptor binding do not correlate with those areas showing a decrease in muscarinic receptor expression. Apparently, the decline in muscarinic acetylcholine receptor density with age does not result from a decline in receptor gene expression.

Tolerance to nicotine-induced sympathoadrenal stimulation and cross-tolerance to stress: differential central and peripheral mechanisms in rats.

Nicotine stimulates the secretion of catecholamines from sympathetic nerve endings and adrenal medulla by acting on peripheral nicotinic cholinergic receptors. Nicotine is also a potent stimulant in the central nervous system but the significance of nicotinic receptors in brain in mediating cardiovascular and sympathoadrenal responses to nicotine is unclear. The responses of resting plasma catecholamines, blood pressure and heart rate were compared in rats receiving nicotine, administered either systemically or intracerebroventricularly (i.c.v.). Sympathoadrenal stress responses were also studied in rats rendered tolerant to nicotine from repeated systemic or intraventricular injections. Nicotine, given either intraventricularly or systemically, produced dose-related increases in the concentration of epinephrine in plasma. Little effect on norepinephrine in plasma was observed with nicotine given intraventricularly, indicating predominant stimulation of adrenomedullary pathways. In contrast, nicotine, given systemically, produced comparable increases in both epinephrine and norepinephrine. Blood pressure increased and heart rate fell in response to either intraventricular or systemic administration of nicotine. Rats exhibited tolerance to nicotine 24 hr after a single intraventricular injection; however, tolerance was not detected with systemically injected nicotine unless the injections were given at least every 30 min. Whereas rats rendered tolerant to systemic administration of nicotine were cross-tolerant to stress, with respect to sympathoadrenal stimulation, cross-tolerance with stress was not detected in rats treated with nicotine repeatedly by the intraventricular route. These results indicate that nicotinic receptors in brain modulate the central sympathetic outflow and adapt readily to nicotine stimulation with prolonged tolerance, but are probably not involved in sympathoadrenal stress responses. Peripheral nicotinic receptors, regulating sympathoadrenal secretion of catecholamines, displayed much shorter-lasting tolerance.

Cellular and subcellular immunolocalization of L-glutamate decarboxylase in rat pancreatic islets.

The cellular and subcellular distribution of L-glutamate decarboxylase (GAD), the biosynthetic enzyme for gamma-aminobutyric acid (GABA), was determined immunohistochemically in rat pancreatic islet using light and electron microscopic techniques. The cellular distribution of GAD was determined at the light microscopic level using an elution/re-staining protocol and a computerized digital image processing technique. At this level of resolution, immunofluorescent GAD was observed to be co-localized with immunofluorescent insulin in the islet B-cells and absent in both the A-cells, which contained glucagon, and the D-cells, which contained somatostatin. Subcellular localization of GAD was determined using an electron microscopic, colloidal gold post-embedding protocol and was compared to insulin immunoreactivity in serial sections of the same B-cell. In the same islet B-cell, GAD immunoreactivity appeared predominantly in the extragranular cytoplasm, whereas insulin immunoreactivity was associated with the secretory granules. Quantitative analysis of GAD immunoreactivity in the B-cell revealed 15.3 +/- 1.8 gold particles/micron2 in the cytoplasm, 1.7 +/- 0.2 gold particles/micron2 in the secretory granules, and 0.4 +/- 0.4 gold particles/micron2 in the mitochondria. The results of this study, localization of the biosynthetic enzyme for GABA to the B-cell cytoplasmic compartment and its absence in the secretory granules which contain insulin, are compatible with the hypothesis that GABA functions as an intracellular mediator of B-cell activity.

A method for immunofluorescent demonstration of three coexisting neurotransmitters in rat brain and spinal cord, using the fluorophores fluorescein, lissamine rhodamine, and 7-amino-4-methylcoumarin-3-acetic acid.

Coexistence of neurotransmitters within single nerve fibers or terminals can be convincingly demonstrated by the use of multicolor immunofluorescence. The present study examined whether three-color immunocytochemical localization of coexisting neurotransmitters can be performed using the blue fluorophore AMCA. Spectrofluorometric examination of secondary antibodies conjugated with AMCA, fluorescein, and lissamine rhodamine showed that the peaks of excitation and emission were well separated and that dots of AMCA-conjugated IgG dried on slides were not visible when viewed using microscope filters for rhodamine and fluorescein. These findings suggest that AMCA might be suitable for three-color immunofluorescence. The usefulness of AMCA for triple labeling was tested directly by staining sections of rat brainstem and spinal cord for serotonin (5HT), substance P (SP), and either enkephalin (ENK) or prepro-thyrotropin-releasing hormone 160-169 (ppT), a marker peptide for thyrotropin-releasing hormone. Triple labeling for 5HT, SP, and ppT was observed in both brainstem and spinal cord but was only very rarely observed for 5HT,SP, and ENK. No evidence was found for artifactual triple labeling, although false negatives appeared to be possible in some circumstances. We conclude that AMCA can be combined with fluorescein and lissamine rhodamine for three-color immunofluorescent studies of coexisting neurotransmitters. In addition, the coexistence of 5HT with ENK appears to be much less common than the coexistence of 5HT with either SP or ppT.

Receptor pharmacology of MDMA and related hallucinogens.

The data presented herein appear to strongly implicate the brain 5HT2 receptor as the site-of-action of the hallucinogenic PIAs and LSD. If so, this discovery represents a major step in understanding the molecular pharmacology of hallucinogenic drugs. Using radioactive hallucinogenic drugs, detailed properties of brain 5HT2 receptors indicating the interaction of 5HT2 receptors with GTP-binding proteins have been revealed. Autoradiographic studies have revealed an extensive cortical distribution of brain 5HT2 receptors; these studies have also suggested that the PIAs may be 5HT1C agonists. Radiolabeling studies in conjunction with drug discrimination studies indicate that MDMA is apparently "amphetamine-like" and not "LSD-like" while MDA is apparently both "LSD-like" and "amphetamine-like." However, MDMA does appear to possess the potential to act as a 5HT2 agonist at high dosages.

Activation of angiotensin II receptors in brain potentiates the stimulating effect of endogenous opioid neurons on central sympathetic outflow.

Endogenous opioid peptides appear to have neurotransmitter or neuromodulator functions in brain mediating a wide variety of effects. We have reported that intracisternal administration of synthetic human beta-endorphin increases plasma concentration of catecholamines, apparently by acting at unknown brain sites to increase sympathetic outflow to the adrenal medulla and sympathetic nerves. In the present study we examined the possibility that angiotensin II, acting in brain, modulates endorphin-induced catecholamine secretion. Simultaneous intracisternal administration of angiotensin II 1.0 nmol together with synthetic human beta-endorphin 1.45 nmol potentiated the plasma epinephrine, norepinephrine and dopamine responses to intracisternal beta-endorphin. In contrast, simultaneous intracisternal administration of the angiotensin II antagonist, [Sar1, Val5, Ala8]-angiotensin II (saralasin), 1.1 nmol together with beta-endorphin, blunted the plasma epinephrine, norepinephrine and dopamine responses to beta-endorphin. These data are consistent with the hypothesis that activation of angiotensin II receptors in brain potentiates the endorphin-induced stimulation of central sympathetic outflow. It remains to be demonstrated whether angiotensin II acting in brain to modulate activity of opioid neurons is synthesized in brain or is derived peripherally.

Hypothalamic opioid peptide regulation of catecholamine secretion.

We compared the plasma catecholamine responses to intracisternal beta-endorphin with those to two mu receptor agonists. Morphine was less potent and [D-Ala2, MePhe4, Gly5-ol]enkephalin (DAGO) was more potent in stimulating catecholamine secretion. Since the hypothalamic paraventricular nucleus (PVN) contains a high level of opioid binding sites and is important for sympathoadrenal regulation, we examined the effects on catecholamine secretion of DAGO infusion into the PVN. DAGO infused into the PVN produced dose-related increases in plasma catecholamine concentrations, with an effective dose as low as 10pmol. This DAGO effect was blocked by the prior systemic administration of naloxone, and DAGO was ineffective when infused into frontoparietal cortex. Thus, endogenous opioid peptides appear to increase central sympathetic outflow and catecholamine secretion by stimulating mu receptors in the PVN.

beta-Endorphin-induced hyperglycemia is mediated by increased central sympathetic outflow to adrenal medulla.

Synthetic human beta-endorphin increased plasma glucose concentration when administered intracisternally in chronically cannulated, conscious, unrestrained, adult male rats. This hyperglycemic effect of beta-endorphin was blocked by prior systemic administration of naloxone, supporting mediation of the effect at opioid receptors in brain. Adrenal denervation blocked the beta-endorphin-induced increase in plasma glucose, supporting a thesis that this effect is mediated at least in part by increased epinephrine secretion. The hyperglycemic response to intracerebral beta-endorphin was also blocked by either intracerebral hemicholinium-3 or somatostatin, supporting both a cholinergic link and a somatostatin neuron in the brain mechanism regulating endorphin-induced stimulation of sympathetic outflow.

Thyrotropin-releasing hormone in spinal cord: coexistence with serotonin and with substance P in fibers and terminals apposing identified preganglionic sympathetic neurons.

In this study we utilized the technique of simultaneous immunofluorescent double-labeling to investigate possible coexistence of the putative neurotransmitter thyrotropin-releasing hormone (TRH) with serotonin (5-HT) and with substance P (SP) in the intermediolateral cell column (IML) of rat spinal cord. We observed fibers and terminals immunoreactive for both TRH and 5-HT or TRH and SP in IML. In addition, this technique was used in animals in which we retrogradely labeled, with fluorescent tracer dyes, preganglionic sympathetic neurons within IML from either the adrenal medulla or the proximal cut end of the cervical sympathetic trunk. In these animals, fibers and terminals containing these combinations of neurotransmitters appeared to oppose identified preganglionic sympathetic neurons in IML. These data represent the first direct immunohistochemical demonstration of fibers and terminals in spinal cord which display coexistence of TRH- with either 5-HT- or SP-immunoreactivity. In addition, the proximity of TRH-immunoreactive fibers and terminals to sympathetic preganglionic neurons in IML support a role for TRH in the regulation of central sympathetic outflow.

Mu receptors at discrete hypothalamic and brainstem sites mediate opioid peptide-induced increases in central sympathetic outflow.

Synthetic human beta-endorphin, 7.25 nmol intracisternally, in conscious, freely moving, cannulated adult male rats increased plasma concentrations of the 3 catecholamines, epinephrine, norepinephrine and dopamine. Similarly administered equimolar morphine increased only plasma epinephrine concentration significantly. A 10-fold greater intracisternal dose of morphine significantly increased plasma concentrations of all 3 catecholamines. This effect was inhibited by prior intra-arterial naloxone administration. Intracisternal administration of the selective mu receptor agonist [D-Ala2,NMe-Phe4,Gly-ol5]enkephalin (DAGO), 2.9 nmol, also increased plasma concentrations of the 3 catecholamines and, furthermore, these effects were significantly greater than those noted in response to equimolar beta-endorphin. The greater potency of DAGO than beta-endorphin to increase catecholamine secretion suggests that this opioid peptide-induced effect is mediated at mu receptors. Administration of DAGO, 0.1 nmol, directly into either the hypothalamic paraventricular nucleus (PVN) or brainstem nucleus of the solitary tract (NTS) significantly increased plasma concentrations of all 3 catecholamines when compared with either saline-infused controls or animals administered DAGO into other brain areas. These catecholamine-stimulating effects of DAGO administered into either PVN or NTS were prevented by prior intra-arterial naloxone administration. Heart rate, but not mean arterial blood pressure, increased in response to DAGO administration into the NTS while no significant cardiovascular changes were noted among the experimental groups in response to DAGO administered into the PVN. These data support a hypothesis that mu receptors at discrete and anatomically distant brain sites mediate opioid peptide-induced catecholamine secretion through activation of the central sympathetic outflow to the adrenal medulla and sympathetic nerve terminals.

Beta-endorphin-induced increases in plasma epinephrine, norepinephrine and dopamine in rats: inhibition of adrenomedullary response by intracerebral somatostatin.

Synthetic human beta-endorphin, 7.25 nmol intracisternally, in unanesthetized, freely moving, chronically cannulated, adult male rats increased plasma concentrations of all 3 catecholamines: epinephrine, norepinephrine and dopamine, for the 2 h period studied. Blockade of these endorphin effects by the prior systemic administration of naloxone supports mediation of the effects at opioid receptors. Acute systemic administration of guanethidine, which decreases norepinephrine release induced by sympathetic nerve stimulation, blunted the plasma norepinephrine response to intracerebral beta-endorphin. Thus, it seems likely that in addition to secretion by adrenal medulla a considerable portion of the beta-endorphin-induced increase in norepinephrine is derived from sympathetic nerve endings. Simultaneous intracisternal administration of another neuropeptide, somatostatin, together with beta-endorphin markedly inhibited the plasma epinephrine response to beta-endorphin, while decreasing the dopamine and norepinephrine responses to a much lesser degree. The dats suggest that beta-endorphin stimulates central sympathetic outflow to both adrenal medulla and sympathetic nerve endings, and further that somatostatin inhibits the effect of endorphin to stimulate outflow to adrenal medulla but does not affect outflow to sympathetic nerve endings.

Fatty acid incorporation depicts brain activity in a rat model of Parkinson's disease.

Following pulse labeling with [3H]arachidonic acid ([3H]AA), its incorporation pattern in brain reflects regional changes in neurotransmitter signal transduction using phospholipase A2, that is, functional activity. In a rat model of Parkinson's disease, unilateral 6-hydroxydopamine lesion in the substantia nigra, [3H]AA acid incorporation from blood was increased in cerebral cortex, caudate putamen, globus pallidus, entopeduncular nucleus, subthalamic nucleus and substantia nigra pars reticulata ipsilateral to the lesion. This increased [3H]AA incorporation likely reflects disinhibition of basal ganglia and cortical circuits secondary to absent inhibitory nigrostriatal dopaminergic input.

Haloperidol downregulates phospholipase A(2) signaling in rat basal ganglia circuits.

Our laboratory has developed an in vivo method to quantitatively evaluate phospholipase A(2) (PLA(2))-mediated signal transduction in brain regions of rodents. In this method, quantitative autoradiography is used to identify brain uptake of intravenously injected, radiolabeled arachidonic acid ([3H]AA). Dopamine D(2) receptors are coupled to G-proteins that activate PLA(2), releasing AA from the stereospecifically numbered (sn) 2 position of phospholipids, and regional [3H]AA uptake is proportional to the rate of release. In the present experiment, the D(2) antagonist haloperidol (1.0 mg/kg i.p.) or the drug vehicle was administered to male adult rats for 21 days. Rats were infused 3 days later with 1.75 mCi/kg [3H]AA (i.v.), anesthetized and decapitated 20 min after infusion onset, and brains were processed for quantitative autoradiography. Chronic haloperidol significantly decreased [3H]AA incorporation in two primary dopaminergic basal ganglia-frontal cortex circuits, the mesocorticolimbic and nigrostriatal systems, while insignificant changes in AA incorporation were noted in other brain regions. These results suggest that one mechanism by which haloperidol exerts its effect is by downregulating D(2)-mediated PLA(2) signaling involving AA release in basal ganglia-frontal cortex circuitry.

Sequelae of parenteral domoic acid administration in rats: comparison of effects on different metabolic markers in brain.

Parenterally administered domoic acid, a structural analog of the excitatory amino acids glutamic acid and kainic acid, has specific effects on brain histology in rats, as measured using different anatomic markers. Domoic acid-induced convulsions affects limbic structures such as hippocampus and entorhinal cortex, and different anatomic markers can detect these neurotoxic effects to varying degrees. Here we report effects of domoic acid administration on quantitative indicators of brain metabolism and gliosis. Domoic acid, 2.25 mg/kg i.p., caused stereotyped behavior and convulsions in approximately 60% of rats which received it. Six to eight days after domoic acid or vehicle administration, the animals were processed to measure regional brain incorporation of the long-chain fatty acids [1-(14)C]arachidonic acid ([14C]AA) and [9,10-(3)H]palmitic acid ([3H]PA), or regional cerebral glucose utilization (rCMRglc) using 2-[1-(14)C]deoxy-D-glucose, by quantitative autoradiography. Others rats were processed to measure brain glial fibrillary acidic protein (GFAP) by enzyme-linked immunosorbent assay. Domoic acid increased GFAP in the anterior portion of cerebral cortex, the caudate putamen and thalamus compared with vehicle. However, in rats that convulsed after domoic acid GFAP was significantly increased throughout the cerebral cortex, as well as in the hippocampus, septum, caudate putamen, and thalamus. Domoic acid, in the absence of convulsions, decreased relative [14C]AA incorporation in the claustrum and pyramidal cell layer of the hippocampus compared with vehicle-injected controls. In the presence of convulsions, relative [14C]AA incorporation was decreased in hippocampus regions CA1 and CA2. Uptake of [3H]PA into brain was unaffected. Relative rCMRglc decreased in entorhinal cortex following domoic acid administration with or without convulsions. These results suggest that acute domoic acid exposure affects discrete brain circuits by inducing convulsions, and that domoic acid-induced convulsions cause chronic effects on brain function that are reflected in altered fatty acid metabolism and gliosis.

Differential identification and localization of adenylyl cyclase and glucose transporter in brain using iodinated derivatives of forskolin.

Two radioiodinated derivatives of forskolin, [125I]6-IHPP-Fsk and [125I]7-IHPP-Fsk, were synthesized as specific ligands for adenylyl cyclase and glucose transporter, respectively. [125I]6-IHPP-Fsk bound to bovine brain homogenates with a Kd of 9 nM and binding was inhibited by forskolin but not 1,9-dideoxyforskolin, cytochalasin B, or D-glucose. [125I]7-IHPP-Fsk bound to bovine brain homogenates at two classes of binding sites with Kd's of 56 nM and 4.7 microM; cytochalasin B and D-glucose inhibited 75% of the high affinity binding while having no effect on the low affinity binding. [125I]6-IHPP-Fsk and [125I]7-IHPP-Fsk were used to localize adenylyl cyclase and glucose transporter in rat brain by receptor autoradiography. The pattern of binding obtained with [125I]6-IHPP-Fsk was similar to that observed using [3H]forskolin to detect adenylyl cyclase. In contrast, the pattern of binding obtained with [125I]7-IHPP-Fsk was similar to that observed by others using [3H]cytochalasin B to detect glucose transporter. These iodinated ligands are selective for adenylyl cyclase and glucose transporter and require significantly shorter exposure times to yield autoradiographs than tritiated ligands.

Simultaneous observation of fluorescent retrogradely labeled neurons and the immunofluorescently labeled fibers apposing them using Fluoro-Gold and antisera labeled with the blue fluorochrome 7-amino-4-methylcoumarin-3-acetic acid (AMCA).

In this paper we describe a method allowing the simultaneous observation of fluorescent retrogradely labeled neurons and the immunofluorescently labeled fibers which appose them. This technique employs the fluorescent dye Fluoro-Gold to label neurons retrogradely, and 7-amino-4-methylcoumarin-3-acetic-acid (AMCA) conjugated to a secondary antiserum as a label for fluorescence immunohistochemistry. Both Fluoro-Gold and AMCA are excited by near-ultraviolet (UV) light, but under UV excitation Fluoro-Gold appears yellow and AMCA appears blue. Thus the two fluorochromes are both visible under a single condition of illumination and can be readily distinguished by color. This allows quick and accurate determination of whether or not immunofluorescent fibers appose retrogradely labeled neurons.

Coexistence of serotonin- and substance P-like immunoreactivity in nerve fibers apposing identified sympathoadrenal preganglionic neurons in rat intermediolateral cell column.

In this study we examined the possibility that serotonin (5-HT) and substance P (SP) coexist in fibers and terminals afferent to sympathoadrenal preganglionic (SAP) neurons in the intermediolateral cell column (IML) of the spinal cord. SAP neurons in the IML were identified by retrograde labeling with either Fast Blue or True Blue injected into the adrenal medulla of rats. A simultaneous immunofluorescent double labeling technique was used to identify both 5-HT- and SP-like immunoreactivity in single tissue sections. Labeled SAP neurons were observed which were apposed by fibers immunoreactive for either neurotransmitter, as well as SAP neurons apposed by neither 5-HT- nor SP-like immunoreactive structures. In addition, 5-HT- and SP-like immunoreactivity were observed in separate fibers apposing the same labeled neuron and coexisting in fibers and terminal appearing in apposition to labeled SAP neurons. These data suggest a complex interaction by these neurotransmitters in regulating sympathetic outflow and may provide a model for interpreting conflicting observations concerning the effects of local 5-HT administration on sympathetic nerve activity.

Infrared Spectroscopic Imaging of the Biochemical Modifications Induced in the Cerebellum of the Niemann-Pick type C Mouse.

We have applied Fourier transform infrared (IR) spectroscopic imaging to the investigation of the neuropathologic effects of a genetic lipid storage disease, Niemann-Pick type C (NPC). Tissue sections both from the cerebella of a strain of BALB/c mice that demonstrated morphology and pathology of the human disease and from control animals were used. These samples were analyzed by standard histopathological procedures as well as this new IR imaging approach. The IR absorbance images exhibit contrast based on biochemical variations and allow for the identification of the cellular layers within the tissue samples. Furthermore, these images provide a qualitative description of the localized biochemical differences existing between the diseased and control tissue in the absence of histological staining. Statistical analyses of the IR spectra extracted from individual cell layers of the imaging data sets provide concise quantitative descriptions of these biochemical changes. The results indicate that lipid is depleted specifically in the white matter of the NPC mouse in comparison to the control samples. Minor differences were noted for the granular layers, but no significant differences were observed in the molecular layers of the cerebellar tissue. These changes are consistent with significant demyelination within the cerebellum of the NPC mouse. © 1999 Society of Photo-Optical Instrumentation Engineers.