Genetic dissection of dopaminergic and noradrenergic contributions to catecholaminergic tracts in early larval zebrafish

The catecholamines dopamine and noradrenaline provide some of the major neuromodulatory systems with far-ranging projections in the brain and spinal cord of vertebrates. However, development of these complex systems is only partially understood. Zebrafish provide an excellent model for genetic analysis of neuronal specification and axonal projections in vertebrates. Here, we analyze the ontogeny of the catecholaminergic projections in zebrafish embryos and larvae up to the fifth day of development and establish the basic scaffold of catecholaminergic connectivity. The earliest dopaminergic diencephalospinal projections do not navigate along the zebrafish primary neuron axonal scaffold but establish their own tracts at defined ventrolateral positions. By using genetic tools, we study quantitative and qualitative contributions of noradrenergic and defined dopaminergic groups to the catecholaminergic scaffold. Suppression of Tfap2a activity allows us to eliminate noradrenergic contributions, and depletion of Otp activity deletes mammalian A11-like Otp-dependent ventral diencephalic dopaminergic groups. This analysis reveals a predominant contribution of Otp-dependent dopaminergic neurons to diencephalospinal as well as hypothalamic catecholaminergic tracts. In contrast, noradrenergic projections make only a minor contribution to hindbrain and spinal catecholaminergic tracts. Furthermore, we can demonstrate that, in zebrafish larvae, ascending catecholaminergic projections to the telencephalon are generated exclusively by Otp-dependent diencephalic dopaminergic neurons as well as by hindbrain noradrenergic groups. Our data reveal the Otp-dependent A11-type dopaminergic neurons as the by far most prominent dopaminergic system in larval zebrafish. These findings are consistent with a hypothesis that Otp-dependent dopaminergic neurons establish the major modulatory system for somatomotor and somatosensory circuits in larval fish. J. Comp. Neurol. 518:439–458, 2010. © 2009 Wiley-Liss, Inc

Determining the neurotransmitter concentration profile at active synapses

Establishing the temporal and concentration profiles of neurotransmitters during synaptic release is an essential step towards understanding the basic properties of inter-neuronal communication in the central nervous system. A variety of ingenious attempts has been made to gain insights into this process, but the general inaccessibility of central synapses, intrinsic limitations of the techniques used, and natural variety of different synaptic environments have hindered a comprehensive description of this fundamental phenomenon. Here, we describe a number of experimental and theoretical findings that has been instrumental for advancing our knowledge of various features of neurotransmitter release, as well as newly developed tools that could overcome some limits of traditional pharmacological approaches and bring new impetus to the description of the complex mechanisms of synaptic transmission

Lifespan pigmentation changes of the substantia nigra detected by neuromelanin-sensitive MRI

Background: Neuromelanin is a pigment with strong iron-chelating properties preferentially found in dopaminergic neurons of the substantia nigra, pars compacta (SNpc). Parkinson's disease is characterized by pronounced, MRI detectable neuromelanin loss, but the neuroprotective or neurotoxic role of neuromelanin remains debated. Histological studies demonstrated neuromelanin increases with age, but this has not been confirmed in vivo and there is uncertainty whether neuromelanin declines, stabilizes or increases from middle age.Methods: This study aimed to establish physiological changes of pigmentation of the SNpc using a pooled dataset of neuromelanin-sensitive 3T MRI from 134 healthy individuals, aged 5~83 years. Neuromelanin-related brightness (regional contrast-to-ratio) and calibrated hyperintense volumes were analyzed using linear and non-linear regression models to characterize age effects. Laterality, sex, and subregional effects were also assessed.Results: For brightness, age effects were best described as a quadratic trajectory explaining 81.5% of the observed variance in SNpc showing a strong increase from childhood to adolescence, with plateauing in middle age and a decline in older age. Similar but less pronounced effects were seen in hyperintense volumes. We also show an anteriorposterior gradient in SNpc contrast, larger normalized NM-rich volume in women >47 of age, but no laterality effect.Conclusions: Using optimized neuromelanin-MRI in a lifespan sample, we demonstrate a strong age effect with an inverted U-shape of SNpc pigmentation-related contrast from childhood to old age. This age trajectory of physiological SNpc pigmentation needs to be taken into account for diagnostic applications of depigmentation. The study also paves the way for systematic investigations of the mechanisms of neuromelanin in healthy and pathological brain development and aging

Measurement of the Bottom-Strange Meson Mixing Phase in the Full CDF Data Set

We report a measurement of the bottom-strange meson mixing phase \beta_s using the time evolution of B0_s -> J/\psi (->\mu+\mu-) \phi (-> K+ K-) decays in which the quark-flavor content of the bottom-strange meson is identified at production. This measurement uses the full data set of proton-antiproton collisions at sqrt(s)= 1.96 TeV collected by the Collider Detector experiment at the Fermilab Tevatron, corresponding to 9.6 fb-1 of integrated luminosity. We report confidence regions in the two-dimensional space of \beta_s and the B0_s decay-width difference \Delta\Gamma_s, and measure \beta_s in [-\pi/2, -1.51] U [-0.06, 0.30] U [1.26, \pi/2] at the 68% confidence level, in agreement with the standard model expectation. Assuming the standard model value of \beta_s, we also determine \Delta\Gamma_s = 0.068 +- 0.026 (stat) +- 0.009 (syst) ps-1 and the mean B0_s lifetime, \tau_s = 1.528 +- 0.019 (stat) +- 0.009 (syst) ps, which are consistent and competitive with determinations by other experiments.Comment: 8 pages, 2 figures, Phys. Rev. Lett 109, 171802 (2012

Dopamine acting at D1-like, D2-like and α1-adrenergic receptors differentially modulates theta and gamma oscillatory activity in primary motor cortex

The loss of dopamine (DA) in Parkinson’s is accompanied by the emergence of exaggerated theta and beta frequency neuronal oscillatory activity in the primary motor cortex (M1) and basal ganglia. DA replacement therapy or deep brain stimulation reduces the power of these oscillations and this is coincident with an improvement in motor performance implying a causal relationship. Here we provide in vitro evidence for the differential modulation of theta and gamma activity in M1 by DA acting at receptors exhibiting conventional and non-conventional DA pharmacology. Recording local field potentials in deep layer V of rat M1, co-application of carbachol (CCh, 5 μM) and kainic acid (KA, 150 nM) elicited simultaneous oscillations at a frequency of 6.49 ± 0.18 Hz (theta, n = 84) and 34.97 ± 0.39 Hz (gamma, n = 84). Bath application of DA resulted in a decrease in gamma power with no change in theta power. However, application of either the D1-like receptor agonist SKF38393 or the D2-like agonist quinpirole increased the power of both theta and gamma suggesting that the DA-mediated inhibition of oscillatory power is by action at other sites other than classical DA receptors. Application of amphetamine, which promotes endogenous amine neurotransmitter release, or the adrenergic α1-selective agonist phenylephrine mimicked the action of DA and reduced gamma power, a result unaffected by prior co-application of D1 and D2 receptor antagonists SCH23390 and sulpiride. Finally, application of the α1-adrenergic receptor antagonist prazosin blocked the action of DA on gamma power suggestive of interaction between α1 and DA receptors. These results show that DA mediates complex actions acting at dopamine D1-like and D2-like receptors, α1 adrenergic receptors and possibly DA/α1 heteromultimeric receptors to differentially modulate theta and gamma activity in M1

Cholinergic Interneurons Mediate Fast VGluT3-Dependent Glutamatergic Transmission in the Striatum

The neurotransmitter glutamate is released by excitatory projection neurons throughout the brain. However, non-glutamatergic cells, including cholinergic and monoaminergic neurons, express markers that suggest that they are also capable of vesicular glutamate release. Striatal cholinergic interneurons (CINs) express the Type-3 vesicular glutamate transporter (VGluT3), although whether they form functional glutamatergic synapses is unclear. To examine this possibility, we utilized mice expressing Cre-recombinase under control of the endogenous choline acetyltransferase locus and conditionally expressed light-activated Channelrhodopsin2 in CINs. Optical stimulation evoked action potentials in CINs and produced postsynaptic responses in medium spiny neurons that were blocked by glutamate receptor antagonists. CIN-mediated glutamatergic responses exhibited a large contribution of NMDA-type glutamate receptors, distinguishing them from corticostriatal inputs. CIN-mediated glutamatergic responses were insensitive to antagonists of acetylcholine receptors and were not seen in mice lacking VGluT3. Our results indicate that CINs are capable of mediating fast glutamatergic transmission, suggesting a new role for these cells in regulating striatal activity

Human SOD2 Modification by Dopamine Quinones Affects Enzymatic Activity by Promoting Its Aggregation: Possible Implications for Parkinson’s Disease

Mitochondrial dysfunction and oxidative stress are considered central in dopaminergic neurodegeneration in Parkinson’s disease (PD). Oxidative stress occurs when the endogenous antioxidant systems are overcome by the generation of reactive oxygen species (ROS). A plausible source of oxidative stress, which could account for the selective degeneration of dopaminergic neurons, is the redox chemistry of dopamine (DA) and leads to the formation of ROS and reactive dopamine-quinones (DAQs). Superoxide dismutase 2 (SOD2) is a mitochondrial enzyme that converts superoxide radicals to molecular oxygen and hydrogen peroxide, providing a first line of defense against ROS. We investigated the possible interplay between DA and SOD2 in the pathogenesis of PD using enzymatic essays, site-specific mutagenesis, and optical and high-field-cw-EPR spectroscopies. Using radioactive DA, we demonstrated that SOD2 is a target of DAQs. Exposure to micromolar DAQ concentrations induces a loss of up to 50% of SOD2 enzymatic activity in a dose-dependent manner, which is correlated to the concomitant formation of protein aggregates, while the coordination geometry of the active site appears unaffected by DAQ modifications. Our findings support a model in which DAQ-mediated SOD2 inactivation increases mitochondrial ROS production, suggesting a link between oxidative stress and mitochondrial dysfunction

Clinical application of biological markers for treatments of resectable non-small-cell lung cancers

We performed a clinical study to identify biological markers useful for the treatment of resectable non-small-cell lung cancers (NSCLCs). In all, 173 patients were studied. By immunohistochemistry, we evaluated the Ki-67 proliferation index, tumour vascularity, thymidylate synthase (TS), vascular endothelial growth factor (VEGF)-A, VEGF-C, and E (epithelial)-cadherin. Concerning the survival of NSCLC patients, tumour vascularity (P<0.01), VEGF-A status (P=0.03), VEGF-C status (P=0.03), and E-cadherin status (P=0.03) were significant prognostic factors in patients with stage I NSCLCs. The Ki-67 proliferation index (P=0.02) and TS status (P<0.01) were significant prognostic factors in patients with stage II–III NSCLCs. In patients with stage II–III NSCLCs, furthermore, the survival of UFT (a combination of tegafur and uracil)-treated patients with TS-negative tumours was significantly better than those of any other patients. Biological markers associated with tumour angiogenesis or metastasis are useful for the detection of aggressive tumours among early-stage NSCLCs. Postoperative chemotherapy might be necessary in such tumours even in stage I. In contrast, tumour proliferation rate and TS status are useful markers for identifying less aggressive tumours in locally advanced NSCLCs. Thymidylate synthase expression is also a useful marker to evaluate responsiveness of UFT-based chemotherapy for these tumours