Structural basis for noradrenergic regulation of neural circuits in the mouse olfactory bulb.

Olfactory input is processed in the glomerulus of the main olfactory bulb (OB) and relayed to higher centers in the brain by projection neurons. Conversely, centrifugal inputs from other brain regions project to the OB. We have previously analyzed centrifugal inputs into the OB from several brain regions using single-neuron labeling. In this study, we analyzed the centrifugal noradrenergic (NA) fibers derived from the locus coeruleus (LC), because their projection pathways and synaptic connections in the OB have not been clarified in detail. We analyzed the NA centrifugal projections by single-neuron labeling and immunoelectron microscopy. Individual NA neurons labeled by viral infection were three-dimensionally traced using Neurolucida software to visualize the projection pathway from the LC to the OB. Also, centrifugal NA fibers were visualized using an antibody for noradrenaline transporter (NET). NET immunoreactive (-ir) fibers contained many varicosities and synaptic vesicles. Furthermore, electron tomography demonstrated that NET-ir fibers formed asymmetrical synapses of varied morphology. Although these synapses were present at varicosities, the density of synapses was relatively low throughout the OB. The maximal density of synapses was found in the external plexiform layer; about 17% of all observed varicosities contained synapses. These results strongly suggest that NA-containing fibers in the OB release NA from both varicosities and synapses to influence the activities of OB neurons. The present study provides a morphological basis for olfactory modulation by centrifugal NA fibers derived from the LC.

Molecular Scaffolds as Double-Targeting Agents For the Diagnosis and Treatment of Neuroblastoma.

The selective delivery of therapeutic and imaging agents to tumoral cells has been postulated as one of the most important challenges in the nanomedicine field. Meta-iodobenzilguanidine (MIBG) is widely used for the diagnosis of neuroblastoma (NB) due to its strong affinity for the norepinephrine transporter (NET), usually overexpressed on the membrane of malignant cells. Herein, a family of novel Y-shaped scaffolds has been synthesized, which have structural analogues of MIBG covalently attached at each end of the Y-structure. The cellular uptake capacity of these double-targeting ligands has been evaluated in vitro and in vivo, yielding one specific Y-shaped structure that is able to be engulfed by the malignant cells, and accumulates in the tumoral tissue, at significantly higher levels than the structure containing only one single targeting agent. This Y-shaped ligand can provide a powerful tool for the current treatment and diagnosis of this disease.

Prevalence and Clinical Correlations of Somatostatin Receptor-2 (SSTR2) Expression in Neuroblastoma.

Alternative radiolabeled, targeted agents are being investigated for children with relapsed neuroblastoma (NB) who do not respond to I-metaiodobenzylguanidine (MIBG) therapy. (DOTA-Tyr)-octreotate targets somatostatin receptors (SSTRs), particularly SSTR2, which are expressed on NB cells. We investigated SSTR2 expression in NB tumors (36 high-risk [HR]; 33 non-HR patients) and correlated SSTR2 levels with clinical features, norepinephrine transporter (NET) expression, and MIBG avidity. SSTR2 and NET immunohistochemistry scores (0 to 3) were calculated on biopsies using digital image analysis based on staining intensity and distribution. Clinical data were correlated with SSTR2 expression. Median SSTR2 score for 69 patients was 1.31 (0.26 to 2.55). Non-HR NB was associated with a higher SSTR2 score (P=0.032). The SSTR2 expression did not correlate with age, International Neuroblastoma Staging System (INSS) stage, MYCN amplification and histology. Higher SSTR2 scores were observed in MIBG-avid versus MIBG-nonavid NB. SSTR2 score was not significantly associated with NET score (r=-0.062, P=0.62). Twenty-six patients who relapsed or progressed had a median SSTR2 score of 1.33 (0.26 to 2.55). Patients with NB including relapsed or progressive disease showed SSTR2 expression at diagnosis, suggesting they could be candidates for radiolabeled-DOTA-conjugated peptide imaging or therapy.

Synthesis, analytical characterization, and monoamine transporter activity of the new psychoactive substance 4-methylphenmetrazine (4-MPM), with differentiation from its ortho- and meta- positional isomers.

The availability of new psychoactive substances (NPS) on the recreational drug market continues to create challenges for scientists in the forensic, clinical and toxicology fields. Phenmetrazine (3-methyl-2-phenylmorpholine) and an array of its analogs form a class of psychostimulants that are well documented in the patent and scientific literature. The present study reports on two phenmetrazine analogs that have been encountered on the NPS market following the introduction of 3-fluorophenmetrazine (3-FPM), namely 4-methylphenmetrazine (4-MPM), and 3-methylphenmetrazine (3-MPM). This study describes the syntheses, analytical characterization, and pharmacological evaluation of the positional isomers of MPM. Analytical characterizations employed various chromatographic, spectroscopic, and mass spectrometric platforms. Pharmacological studies were conducted to assess whether MPM isomers might display stimulant-like effects similar to the parent compound phenmetrazine. The isomers were tested for their ability to inhibit uptake or stimulate release of tritiated substrates at dopamine, norepinephrine and serotonin transporters using in vitro transporter assays in rat brain synaptosomes. The analytical characterization of three vendor samples revealed the presence of 4-MPM in two of the samples and 3-MPM in the third sample, which agreed with the product label. The pharmacological findings suggest that 2-MPM and 3-MPM will exhibit stimulant properties similar to the parent compound phenmetrazine, whereas 4-MPM may display entactogen properties more similar to 3,4-methylenedioxymethamphetamine (MDMA). The combination of test purchases, analytical characterization, targeted organic synthesis, and pharmacological evaluation of NPS and their isomers is an effective approach for the provision of data on these substances as they emerge in the marketplace.

The regulation of corticosteroid receptors in response to chronic social defeat.

Our previous studies demonstrated that chronic social defeat (CSD) up-regulated expression of the serotonin transporter (SERT) and norepinephrine transporter (NET) in the brain, which was mediated by corticosteroid receptors. In the present study we first analyzed the alterations of corticosteroid receptors in different brain regions after the CSD paradigm. The results showed that CSD significantly reduced glucocorticoid receptor (GR) protein levels in the CA1 and dentate gyrus of the hippocampus, as well as in central and basolateral nuclei of the amygdala, which was accompanied by the translocation of GR from cytoplasm to nuclei. CSD also markedly reduced GR mRNA levels and MR immunoreactivity in the CA1, CA3 and dentate gyrus areas of the hippocampus. Conversely, CSD pronouncedly enhanced GR mRNA and protein levels in the dorsal raphe nucleus and locus coeruleus relative to the control. As an extension of our previous studies, in situ hybridization and immunohistochemical staining demonstrated that CSD regimen caused a notable increase of SERT mRNA levels in the dorsal raphe nucleus and increased SERT immunoreactivities in CA1 and CA3 of the hippocampus, as well as those in the basolateral nuclei of the amygdala. Likewise, CSD regimen resulted in an evident enhancement of NET immunoreactivity in the CA1 of the hippocampus and in the basolateral nuclei of the amygdala. Our current findings suggest that GR expressional alterations in response to CSD are complex and brain region-specific, which may correspond to their different functions in these regions.

Effects and Mechanisms of Radiofrequency Ablation of Renal Sympathetic Nerve on Anti-Hypertension in Canine.

BACKGROUND: Radiofrequency ablation of renal sympathetic nerve (RDN) shows effective BP reduction in hypertensive patients while the specific mechanisms remain unclear. OBJECTIVE: We hypothesized that abnormal levels of norepinephrine (NE) and changes in NE-related enzymes and angiotensinconverting enzyme 2 (ACE2), angiotensin (Ang)-(1-7) and Mas receptor mediate the anti-hypertensive effects of RDN. METHODS: Mean values of systolic blood pressure (SBP), diastolic blood pressure (DBP) and mean arterial pressure (MAP) were assessed at baseline and follow-up. Plasma and renal norepinephrine (NE) concentrations were determined using highperformance liquid chromatography with electrochemical detection, and levels of NE-related enzyme and ACE2-Ang(1-7)- Mas were measured using real time PCR, Western blot and immunohistochemistry or Elisa in a hypertensive canine model fed with high-fat diet and treated with RDN. The parameters were also determined in a sham group treated with renal arteriography and a control group fed with normal diet. RESULTS: RDN decreased SBP, DBP, MAP, plasma and renal NE. Compared with the sham group, renal tyrosine hydroxylase (TH) expression was lower and renalase expression was higher in the RDN group. Compared with the control group, renal TH and catechol-o-methyl transferase (COMT) were higher and renalase was lower in the sham group. Moreover, renal ACE2, Ang-(1-7) and Mas levels of the RDN group were higher than those of the sham group, which were lower than those of the control group. CONCLUSION: RDN shows anti-hypertensive effect with reduced NE and activation of ACE2-Ang(1-7)-Mas, indicating that it may contribute to the anti-hypertensive effect of RDN.

Effects and Mechanisms of Radiofrequency Ablation of Renal Sympathetic Nerve on Anti-Hypertension in Canine / Efeitos e Mecanismos da Denervação Simpática Renal por Radiofrequência na Anti-Hipertensão em Cães

Abstract Background: Radiofrequency ablation of renal sympathetic nerve (RDN) shows effective BP reduction in hypertensive patients while the specific mechanisms remain unclear. Objective: We hypothesized that abnormal levels of norepinephrine (NE) and changes in NE-related enzymes and angiotensinconverting enzyme 2 (ACE2), angiotensin (Ang)-(1-7) and Mas receptor mediate the anti-hypertensive effects of RDN. Methods: Mean values of systolic blood pressure (SBP), diastolic blood pressure (DBP) and mean arterial pressure (MAP) were assessed at baseline and follow-up. Plasma and renal norepinephrine (NE) concentrations were determined using highperformance liquid chromatography with electrochemical detection, and levels of NE-related enzyme and ACE2-Ang(1-7)- Mas were measured using real time PCR, Western blot and immunohistochemistry or Elisa in a hypertensive canine model fed with high-fat diet and treated with RDN. The parameters were also determined in a sham group treated with renal arteriography and a control group fed with normal diet. Results: RDN decreased SBP, DBP, MAP, plasma and renal NE. Compared with the sham group, renal tyrosine hydroxylase (TH) expression was lower and renalase expression was higher in the RDN group. Compared with the control group, renal TH and catechol-o-methyl transferase (COMT) were higher and renalase was lower in the sham group. Moreover, renal ACE2, Ang-(1-7) and Mas levels of the RDN group were higher than those of the sham group, which were lower than those of the control group. Conclusion: RDN shows anti-hypertensive effect with reduced NE and activation of ACE2-Ang(1-7)-Mas, indicating that it may contribute to the anti-hypertensive effect of RDN.

Resumo Fundamentos: A denervação simpática renal por radiofrequência (DSR) mostra redução eficaz da pressão arterial (PA) de pacientes hipertensos, ainda que os mecanismos específicos permaneçam obscuros. Objetivo: Fizemos a hipótese de que níveis alterados de noradrenalina (NA) e mudanças nas enzimas relacionadas à NA e enzima conversora de angiotensina 2 (ECA-2), angiotensina (Ang)-(1-7) e receptor Mas são mediadores dos efeitos antihipertensivos da DSR. Métodos: Foram avaliados os valores médios de pressão arterial sistólica (PAS), pressão arterial diastólica (PAD) e pressão arterial média (PAM) no início e durante o seguimento. Foram medidas as concentrações plasmática e renal de noradrenalina (NA) por cromatografia líquida de alta eficiência com detecção eletroquímica, e os níveis de enzima relacionada à NA e ECA2-Ang-(1-7)-Mas através de PCR em tempo real, Western blot e imunohistoquímica ou Elisa em um modelo canino de hipertensão que recebeu ração rica em gordura e foi tratado com DSR. Os parâmetros também foram determinados em um grupo de cirurgia simulada submetido à arteriografia renal e em um grupo controle que recebeu dieta normal. Resultados: DSR causou diminuição da PAS, PAD, PAM e das concentrações plasmática e renal de NA. Em comparação ao grupo placebo, a expressão da tirosina hidroxilase (TH) renal foi menor e a da renalase foi maior no grupo DSR. Em comparação ao grupo controle, os níveis de TH renal e de catecol-o-metil-transferase (COMT) foram maiores e os de renalase foram menores no grupo cirurgia simulada. Além disso, os níveis renais de ECA2, Ang-(1-7) e Mas foram maiores no grupo DSR do que no grupo cirurgia simulada, que, por sua vez, foram menores do que no grupo controle. Conclusões: A DSR mostra efeitos anti-hipertensivos com redução da NA e ativação da ECA2-Ang-(1-7)-Mas, o que indica que pode contribuir com o efeito anti-hipertensivo da DSR.

Differential Internalization Rates and Postendocytic Sorting of the Norepinephrine and Dopamine Transporters Are Controlled by Structural Elements in the N Termini.

The norepinephrine transporter (NET) mediates reuptake of synaptically released norepinephrine in central and peripheral noradrenergic neurons. The molecular processes governing availability of NET in the plasma membrane are poorly understood. Here we use the fluorescent cocaine analogue JHC 1-64, as well as several other approaches, to investigate the trafficking itinerary of NET in live noradrenergic neurons. Confocal imaging revealed extensive constitutive internalization of JHC 1-64-labeled NET in the neuronal somata, proximal extensions and presynaptic boutons. Phorbol 12-myristate 13-acetate increased intracellular accumulation of JHC 1-64-labeled NET and caused a parallel reduction in uptake capacity. Internalized NET strongly colocalized with the "long loop" recycling marker Rab11, whereas less overlap was seen with the "short loop" recycling marker Rab4 and the late endosomal marker Rab7. Moreover, mitigating Rab11 function by overexpression of dominant negative Rab11 impaired NET function. Sorting of NET to the Rab11 recycling compartment was further supported by confocal imaging and reversible biotinylation experiments in transfected differentiated CATH.a cells. In contrast to NET, the dopamine transporter displayed markedly less constitutive internalization and limited sorting to the Rab11 recycling compartment in the differentiated CATH.a cells. Exchange of domains between the two homologous transporters revealed that this difference was determined by non-conserved structural elements in the intracellular N terminus. We conclude that NET displays a distinct trafficking itinerary characterized by continuous shuffling between the plasma membrane and the Rab11 recycling compartment and that the functional integrity of the Rab11 compartment is critical for maintaining proper presynaptic NET function.

Iodine-123 labeled reboxetine analogues for imaging of noradrenaline transporter in brain using single photon emission computed tomography.

Preliminary investigation of the radioiodinated (S,S)-reboxetine analogue, (123) I-INER, in baboons showed this tracer to have promise for imaging the noradrenaline transporter (NAT) using single photon emission computed tomography (SPECT). More recently, the radioiodinated (R,S)-stereoisomer of (123) I-INER, (123) I-NKJ64, has been synthesized and preliminary evaluation in rats has been reported. This article reports the brain distribution and pharmacokinetic properties of (123) I-NKJ64 in baboons and compares results with (123) I-INER data in the same species. SPECT studies were conducted in two ovariectomized adult female baboons using two different protocols: (1) bolus of (123) I-INER or (123) I-NKJ64; and (2) bolus plus constant infusion of (123) I-NKJ64 with reboxetine (2.0 mg/kg) administration at equilibrium. Following bolus injection, both radiotracers rapidly and avidly entered the baboon brain. The regional brain accumulation of (123) I-NKJ64 did not match the known distribution of NAT in baboon brain, contrasting with previous results obtained in rats. Conversely, the regional distribution of (123) I-INER was consistent with known distribution of NAT in baboon brain. No displacement of (123) I-NKJ64 was observed following administration of reboxetine. This contrasts with previous data obtained for (123) I-INER, where 60% of specific binding was displaced by a lower dose of reboxetine. These data suggest that (123) I-NKJ64 may lack affinity and selectivity for NAT in baboon brain and (123) I-INER is the most promising iodinated reboxetine analogue developed to date for in vivo imaging of NAT in brain using SPECT. This study highlights the importance of species differences during radiotracer development and the stereochemical configuration of analogues of reboxetine in vivo.

The norepinephrine transporter (NET) radioligand (S,S)-[18F]FMeNER-D2 shows significant decreases in NET density in the human brain in Alzheimer's disease: a post-mortem autoradiographic study.

Earlier post-mortem histological and autoradiographic studies have indicated a reduction of cell numbers in the locus coeruleus (LC) and a corresponding decrease in norepinephrine transporter (NET) in brains obtained from Alzheimer's disease (AD) patients as compared to age-matched healthy controls. In order to test the hypothesis that the regional decrease of NET is a disease specific biomarker in AD and as such, it can be used in PET imaging studies for diagnostic considerations, regional differences in the density of NET in various anatomical structures were measured in whole hemisphere human brain slices obtained from AD patients and age-matched control subjects in a series of autoradiographic experiments using the novel selective PET radioligand for NET (S,S)-[(18)F]FMeNER-D(2). (S,S)-[(18)F]FMeNER-D(2) appears to be a useful imaging biomarker for quantifying the density of NET in various brain structures, including the LC and the thalamus wherein the highest densities are found in physiological conditions. In AD significant decreases of NET densities can be demonstrated with the radioligand in both structures as compared to age-matched controls. The decreases in AD correlate with the progress of the disease as indicated by Braak grades. As the size of the LC is below the spatial resolution of the PET scanners, but the size of the thalamus can be detected with appropriate spatial accuracy in advanced scanners, the present findings confirm our earlier observations with PET that the in vivo imaging of NET with (S,S)-[(18)F]FMeNER-D(2) in the thalamus is viable. Nevertheless, further studies are warranted to assess the usefulness of such an imaging approach for the early detection of changes in thalamic NET densities as a disease-specific biomarker and the possible use of (S,S)-[(18)F]FMeNER-D(2) as a molecular imaging biomarker in AD.

Design and synthesis of (2R,3S)-iodoreboxetine analogues for SPECT imaging of the noradrenaline transporter.

A stereoselective 10-step synthesis of iodophenoxy analogues of (2R,3S)-reboxetine has been developed with the aim of generating a new SPECT imaging agent for the noradrenaline transporter (NAT). In vitro testing of these compounds against various mono-amine transporters showed an ortho-iodophenoxy analogue to have excellent affinity (K(i) 8.4 nM) and good selectivity for NAT.

Stereoselective synthesis of (2S,3R)- and (2R,3S)-iodoreboxetine; potential SPECT imaging agents for the noradrenaline transporter.

With the aim of developing a new SPECT imaging agent for the noradrenaline transporter, a twelve-step stereoselective synthesis of iodinated analogues of (2S,3R)- and (2R,3S)-reboxetine has been achieved from 4-bromobenzaldehyde. The key steps involve a Sharpless asymmetric epoxidation to establish the stereogenic centres and a copper catalysed aromatic halogen exchange reaction to introduce the key iodine atom. In vitro testing of these compounds using a [(3)H]nisoxetine displacement assay with homogenised rat brain shows both compounds to have significant affinity, with K(i) values of 320.8 nM and 58.2 nM for (2S,3R)- and (2R,3S)-iodoreboxetine respectively.

Validation of a fluorescence-based high-throughput assay for the measurement of neurotransmitter transporter uptake activity.

Pre-synaptic dopamine, norepinephrine and serotonin transporters (DAT, NET and SERT) terminate synaptic catecholamine transmission through reuptake of released neurotransmitter. Common approaches for studying these transporters involve radiolabeled substrates or inhibitors which, however, have several limitations. In this study we have used a novel neurotransmitter transporter uptake assay kit. The assay employs a fluorescent substrate that mimics the biogenic amine neurotransmitters and is taken up by the cell through the specific transporters, resulting in increased fluorescence intensity. In order to validate the assay, a variety of reference and proprietary neurotransmitter transporter ligands from a number of chemical and pharmacological classes were tested. The ability of these compounds to inhibit the selective transporter-mediated uptake demonstrated a similar rank order of potency and IC(50) values close to those obtained in radiolabeled neurotransmitter uptake assays. The described assay enables monitoring of dynamic transport activity of DAT, NET and SERT and is amenable for high-throughput screening and compound characterization.

Imaging the norepinephrine transporter with positron emission tomography: initial human studies with (S,S)-[18F]FMeNER-D2.

INTRODUCTION: (S,S)-[(18)F]FMeNER-D(2) is a recently developed positron emission tomography (PET) ligand for in vivo quantification of norepinephrine transporter. A monkey occupancy study with the radioligand indicated that (S,S)-[(18)F]FMeNER-D(2) can be useful for quantitative PET analysis. In this preliminary study, regional distributions in the living human brain were evaluated. MATERIALS AND METHODS: Brain PET measurements were performed for a total of 255 min after the injection of 188.3 +/- 5.7 MBq of (S,S)-[(18)F]FMeNER-D(2) in four healthy male subjects. Regions of interests were drawn on the thalamus and the caudate in the coregistered MRI/PET images. RESULTS: (S,S)-[(18)F]FMeNER-D(2) displayed good brain penetration and selective retention in regions rich in norepinephrine reuptake sites. The transient peak equilibrium was reached during the PET measurements. The ratios of radioactivity uptake in the thalamus to that in the caudate were 1.50 +/- 0.06 for the time period of 90-255 min. CONCLUSION: The present preliminary investigation indicates that (S,S)-[(18)F]FMeNER-D(2) has suitable characteristics for probing the norepinephrine reuptake system with PET in the human brain.

Development of radioligands for imaging of brain norepinephrine transporters in vivo with positron emission tomography.

In the central nervous system (CNS) and in the periphery, specific proteins (transporters) are responsible for the regulation of the synaptic concentrations of the major monoamine neurotransmitters, noradrenaline (NE), serotonin (5-HT) and dopamine (DA). Several reports have shown that the expression of these transporters within the CNS may be altered in patients with certain neurodegenerative or neuropsychiatric disorders. Therefore, in the CNS the monoamine transporters are major targets for existing and developmental drugs. The best known drugs targeting these transporters are the selective 5-HT reuptake inhibitors (SSRIs) (e.g. citalopram, Celexa) that are most frequently used in the treatment of clinical depression. Selective NE reuptake inhibitors (NRIs) have also found use for the treatment of depression and other conditions such as attention deficit hyperactivity (ADHD) disorder. Given that the NE transporter (NET) is also a binding site for cocaine and drugs of abuse, there is a great need for a probe to assess the densities of NET in vivo by brain imaging with either positron emission tomography (PET) or single photon emission tomography (SPET). PET in particular has the potential to measure NET densities quantitatively and with high resolution in the human brain in vivo. The quality of a PET image depends crucially on the radioligand used in the emission measurement. Commonly used radionuclides in PET radioligands are carbon 11 (t(1/2) = 20.4 min) and fluorine-18 (t(1/2) = 109.8 min). This review specifically summarizes the present status of the development of (11)C- or (18)F-labeled ligands as tools for imaging NET in brain with PET in support of neuropsychiatric clinical research and drug development.

Synthesis and biological evaluation of 2beta,3alpha-(substituted phenyl)nortropanes as potential norepinephrine transporter imaging agents.

A series of 2beta,3alpha-(substituted phenyl)nortropanes was synthesized and evaluated in vitro for human monoamine transporters. All compounds studied in this series exhibited nanomolar potency for the norepinephrine transporter (NET). Radiolabeling and nonhuman primate microPET brain imaging studies were performed with the most promising compound, [(11)C]1, to determine its utility as a NET imaging agent. Despite high in vitro affinity for the human NET, the high uptake of [(11)C]1 in the caudate and putamen excludes its use as an in vivo PET imaging agent for the NET.

Clinical catecholamine neurochemistry: a legacy of Julius Axelrod.

1. Discoveries, insights, and concepts that Julius Axelrod introduced about the disposition and metabolism of catecholamines provided the scientific basis and spurred the development of clinical catecholamine neurochemistry. 2. Here, we provide examples of this aspect of Axelrod's scientific legacy.

Chronic stress increases the plasmalemmal distribution of the norepinephrine transporter and the coexpression of tyrosine hydroxylase in norepinephrine axons in the prefrontal cortex.

Norepinephrine (NE) potently modulates the cognitive and affective functions of the prefrontal cortex (PFC). Deficits in NE transmission are implicated in psychiatric disorders, and antidepressant drugs that block the NE transporter (NET) effectively treat these conditions. Our initial ultrastructural studies of the rat PFC revealed that most NE axons (85-90%) express NET primarily within the cytoplasm and lack detectable levels of the synthetic enzyme tyrosine hydroxylase (TH). In contrast, the remaining 10-15% of PFC NE axons exhibit predominantly plasmalemmal NET and evident TH immunoreactivity. These unusual characteristics suggest that most PFC NE axons have an unrecognized, latent capacity to enhance the synthesis and recovery of transmitter. In the present study, we used dual-labeling immunocytochemistry and electron microscopy to examine whether chronic cold stress, a paradigm that persistently increases NE activity, would trigger cellular changes consistent with this hypothesis. After chronic stress, neither the number of profiles exhibiting NET labeling nor their size was changed. However, the proportion of plasmalemmal NET nearly doubled from 29% in control animals to 51% in stressed rats. Moreover, the expression of detectable TH in NET-labeled axons increased from only 13% of profiles in control rats to 32% of profiles in stressed animals. Despite the consistency of these findings, the magnitude of the changes varied across individual rats. These data represent the first demonstration of activity-dependent trafficking of NET and expression of TH under physiological conditions and have important implications for understanding the pathophysiology and treatment of stress-related affective disorders.

Compensatory changes in the noradrenergic nervous system in the locus ceruleus and hippocampus of postmortem subjects with Alzheimer's disease and dementia with Lewy bodies.

In Alzheimer's disease (AD), there is a significant loss of locus ceruleus (LC) noradrenergic neurons. However, functional and anatomical evidence indicates that the remaining noradrenergic neurons may be compensating for the loss. Because the noradrenergic system plays an important role in learning and memory, it is important to determine whether compensation occurs in noradrenergic neurons in the LC and hippocampus of subjects with AD or a related dementing disorder, dementia with Lewy bodies (DLB). We observed profound neuronal loss in the LC in AD and DLB subjects with three major changes in the noradrenergic system consistent with compensation: (1) an increase in tyrosine hydroxylase (TH) mRNA expression in the remaining neurons; (2) sprouting of dendrites into peri-LC dendritic zone, as determined by alpha2-adrenoreceptors (ARs) and norepinephrine transporter binding sites; and (3) sprouting of axonal projections to the hippocampus as determined by alpha2-ARs. In AD and DLB subjects, the postsynaptic alpha1-ARs were normal to elevated. Expression of alpha1A- and alpha2A-AR mRNA in the hippocampus of AD and DLB subjects were not altered, but expression of alpha1D- and alpha2C-AR mRNA was significantly reduced in the hippocampus of AD and DLB subjects. Therefore, in AD and DLB subjects, there is compensation occurring in the remaining noradrenergic neurons, but there does appear to be a loss of specific AR in the hippocampus. Because changes in these noradrenergic markers in AD versus DLB subjects were similar (except neuronal loss and the increase in TH mRNA were somewhat greater in DLB subjects), the presence of Lewy bodies in addition to plaques and tangles in DLB subjects does not appear to further affect the noradrenergic compensatory changes.