Polysialic acid is required for dopamine D2 receptor-mediated plasticity involving inhibitory circuits of the rat medial prefrontal cortex

Decreased expression of dopamine D2 receptors (D2R), dysfunction of inhibitory neurotransmission and impairments in the structure and connectivity of neurons in the medial prefrontal cortex (mPFC) are involved in the pathogenesis of schizophrenia and major depression, but the relationship between these changes remains unclear. The polysialylated form of the neural cell adhesion molecule (PSA-NCAM), a plasticity-related molecule, may serve as a link. This molecule is expressed in cortical interneurons and dopamine, via D2R, modulates its expression in parallel to that of proteins related to synapses and inhibitory neurotransmission, suggesting that D2R-targeted antipsychotics/antidepressants may act by affecting the plasticity of mPFC inhibitory circuits. To understand the role of PSA-NCAM in this plasticity, rats were chronically treated with a D2R agonist (PPHT) after cortical PSA depletion. PPHT-induced increases in GAD67 and synaptophysin (SYN) neuropil expression were blocked when PSA was previously removed, indicating a role for PSA-NCAM in this plasticity. The number of PSA-NCAM expressing interneuron somata also increased after PPHT treatment, but the percentages of these cells belonging to different interneuronal subpopulations did not change. Cortical pyramidal neurons did not express PSA-NCAM, but puncta co-expressing this molecule and parvalbumin could be found surrounding their somata. PPHT treatment increased the number of PSA-NCAM and parvalbumin expressing perisomatic puncta, but decreased the percentage of parvalbumin puncta that co-expressed SYN. PSA depletion did not block these effects on the perisomatic region, but increased further the number of parvalbumin expressing puncta and increased the percentage of puncta co-expressing SYN and parvalbumin, suggesting that the polysialylation of NCAM may regulate perisomatic inhibition of mPFC principal neurons. Summarizing, the present results indicate that dopamine acting on D2R influences structural plasticity of mPFC interneurons and point to PSA-NCAM as a key player in this remodeling.     

Femoral artery remodeling after aerobic exercise training without weight loss in women

Background

It is currently unclear whether reductions in adiposity mediate the improvements in vascular health that occur with aerobic exercise. The purpose of this longitudinal study of 13 healthy women (33 ± 4 years old) was to determine whether 14 weeks of aerobic exercise would alter functional measures of vascular health, namely resting aortic pulse wave velocity (aPWV, an index of arterial stiffness), femoral artery diameter (D_FA), and femoral artery blood flow (BF_FA) independent of changes in adiposity.

Methods

Aerobic fitness was assessed as VO₂peak normalized to fat-free mass, and adiposity (percent body fat) was determined by dual energy x-ray absorptiometry. Serum concentrations of proteins associated with risk for cardiovascular disease, including C-reactive protein (CRP), soluble intercellular adhesion molecule-1 (sICAM-1), and leptin, were also measured. Subjects cycled for 50 minutes, 3 times per week.

Results

Aerobic fitness normalized to fat-free mass increased 6% (P = 0.03) whereas adiposity did not change. Resting D_FA increased 12% (P < 0.001) and resting shear rate decreased 28% (P = 0.007). Aortic PWV, and serum sICAM-1, CRP and leptin did not change with training.

Conclusion

Significant reductions in adiposity were not necessary for aerobic exercise training to bring about improvements in aerobic fitness and arterial remodeling. Peripheral arterial remodeling occurred without changes in central arterial stiffness or markers of inflammation.

     

Circadian remodeling of neuronal circuits involved in rhythmic behavior

Clock output pathways are central to convey timing information from the circadian clock to a diversity of physiological systems, ranging from cell-autonomous processes to behavior. While the molecular mechanisms that generate and sustain rhythmicity at the cellular level are well understood, it is unclear how this information is further structured to control specific behavioral outputs. Rhythmic release of pigment dispersing factor (PDF) has been proposed to propagate the time of day information from core pacemaker cells to downstream targets underlying rhythmic locomotor activity. Indeed, such circadian changes in PDF intensity represent the only known mechanism through which the PDF circuit could communicate with its output. Here we describe a novel circadian phenomenon involving extensive remodeling in the axonal terminals of the PDF circuit, which display higher complexity during the day and significantly lower complexity at nighttime, both under daily cycles and constant conditions. In support to its circadian nature, cycling is lost in bona fide clockless mutants. We propose this clock-controlled structural plasticity as a candidate mechanism contributing to the transmission of the information downstream of pacemaker cells.     

听觉系统抑制性神经回路的活动依赖性组构

最近的研究发现,哺乳动物听觉系统中抑制性神经回路的成熟和优化依赖于自发性和经验性的神经活动,神经活动对于抑制性突触的亚细胞定位、抑制性神经回路的拓扑组构,以及耳优势带的形成均至关重要。这些研究可能揭示了脑发育过程中抑制性神经回路构建的一般规律和细胞机制。     

Rapid loss of bone mass and strength in mice after abdominal irradiation

Localized irradiation is a common treatment modality for malignancies in the pelvic-abdominal cavity. We report here on the changes in bone mass and strength in mice 7-14 days after abdominal irradiation. Male C57BL/6 mice of 10-12 weeks of age were given a single-dose (0, 5, 10, 15 or 20 Gy) or fractionated (3 Gy × 2 per day × 7.5 days) X rays to the abdomen and monitored daily for up to 14 days. A decrease in the serum bone formation marker and ex vivo osteoblast differentiation was detected 7 days after a single dose of radiation, with little change in the serum bone resorption marker and ex vivo osteoclast formation. A single dose of radiation elicited a loss of bone mineral density (BMD) within 14 days of irradiation. The BMD loss was up to 4.1% in the whole skeleton, 7.3% in tibia, and 7.7% in the femur. Fractionated abdominal irradiation induced similar extents of BMD loss 10 days after the last fraction: 6.2% in the whole skeleton, 5.1% in tibia, and 13.8% in the femur. The loss of BMD was dependent on radiation dose and was more profound in the trabecula-rich regions of the long bones. Moreover, BMD loss in the total skeleton and the femurs progressed with time. Peak load and stiffness in the mid-shaft tibia from irradiated mice were 11.2-14.2% and 11.5-25.0% lower, respectively, than sham controls tested 7 days after a single-dose abdominal irradiation. Our data demonstrate that abdominal irradiation induces a rapid loss of BMD in the mouse skeleton. These effects are bone type- and region-specific but are independent of radiation fractionation. The radiation-induced abscopal damage to the skeleton is manifested by the deterioration of biomechanical properties of the affected bone.     

Activity-dependent organization of inhibitory circuits: lessons from the auditory system

In contrast to our detailed knowledge about the development and plasticity of excitatory neuronal circuits, little is known about the development of inhibitory circuits. Recent studies from the developing mammalian auditory system have revealed the presence of substantial activity-dependent synaptic reorganization in several inhibitory pathways. These studies importantly shed some new light on the general rules and cellular mechanisms that manage the organization of precise inhibitory circuits in the developing brain.     

Transient loss of dopamine autoreceptor control in the presence of highly potent dopamine agonists

The concentrations of endogenous ligands generally remain in a bounded range around a basal level, a manifestation of control. The dopaminergic system is an excellent example of a control system in which a negative feedback signal is associated with receptor occupancy of a D2-like dopamine autoreceptor. A consequence of the control theory is that autoreceptor occupancy by an agonist results in dopamine levels below the basal, whereas similar stimulation by a dopamine competitive antagonist results in an increase of dopamine to levels above the basal. These consequences of control theory were tested and verified in the rat striatum by infusing graded doses of either the agonist, quinpirole, or the antagonist, sulpiride, into the rat striatum via a microdialysis probe and sampling dopamine and metabolite levels at various times after the start of infusion. Control was maintained even at the very highest doses of these compounds, i.e., striatal dopamine concentration rose in response to the antagonist and fell in response to the agonist. In contrast, administration of each of two high affinity dopamine agonists, 7-OH-DPAT and PPHT showed dose-dependent control only up to certain doses. Above these doses the dopamine concentration actually increased to levels well above basal, an indication of loss of control. These findings suggest that the control of this endogenous ligand does not extend to the very highest levels of autoreceptor occupancy.     

Tracking the expression of excitatory and inhibitory neurotransmission-related proteins and neuroplasticity markers after noise induced hearing loss

Excessive exposure to loud noise can damage the cochlea and create a hearing loss. These pathologies coincide with a range of CNS changes including reorganisation of frequency representation, alterations in the pattern of spontaneous activity and changed expression of excitatory and inhibitory neurotransmitters. Moreover, damage to the cochlea is often accompanied by acoustic disorders such as hyperacusis and tinnitus, suggesting that one or more of these neuronal changes may be involved in these disorders, although the mechanisms remain unknown. We tested the hypothesis that excessive noise exposure increases expression of markers of excitation and plasticity, and decreases expression of inhibitory markers over a 32-day recovery period. Adult rats (n = 25) were monaurally exposed to a loud noise (16 kHz, 1/10(th) octave band pass (115 dB SPL)) for 1-hour, or left as non-exposed controls (n = 5). Animals were euthanased at either 0, 4, 8, 16 or 32 days following acoustic trauma. We used Western Blots to quantify protein levels of GABA(A) receptor subunit α1 (GABA(A)α1), Glutamic-Acid Decarboxylase-67 (GAD-67), N-Methyl-D-Aspartate receptor subunit 2A (NR2A), Calbindin (Calb1) and Growth Associated Protein 43 (GAP-43) in the Auditory Cortex (AC), Inferior Colliculus (IC) and Dorsal Cochlear Nucleus (DCN). Compared to sham-exposed controls, noise-exposed animals had significantly (p<0.05): lower levels of GABA(A)α1 in the contralateral AC at day-16 and day-32, lower levels of GAD-67 in the ipsilateral DCN at day-4, lower levels of Calb1 in the ipsilateral DCN at day-0, lower levels of GABA(A)α1 in the ipsilateral AC at day-4 and day-32. GAP-43 was reduced in the ipsilateral AC for the duration of the experiment. These complex fluctuations in protein expression suggests that for at least a month following acoustic trauma the auditory system is adapting to a new pattern of sensory input.     

Rapid construction of insulated genetic circuits via synthetic sequence-guided isothermal assembly

In vitro recombination methods have enabled one-step construction of large DNA sequences from multiple parts. Although synthetic biological circuits can in principle be assembled in the same fashion, they typically contain repeated sequence elements such as standard promoters and terminators that interfere with homologous recombination. Here we use a computational approach to design synthetic, biologically inactive unique nucleotide sequences (UNSes) that facilitate accurate ordered assembly. Importantly, our designed UNSes make it possible to assemble parts with repeated terminator and insulator sequences, and thereby create insulated functional genetic circuits in bacteria and mammalian cells. Using UNS-guided assembly to construct repeating promoter-gene-terminator parts, we systematically varied gene expression to optimize production of a deoxychromoviridans biosynthetic pathway in Escherichia coli. We then used this system to construct complex eukaryotic AND-logic gates for genomic integration into embryonic stem cells. Construction was performed by using a standardized series of UNS-bearing BioBrick-compatible vectors, which enable modular assembly and facilitate reuse of individual parts. UNS-guided isothermal assembly is broadly applicable to the construction and optimization of genetic circuits and particularly those requiring tight insulation, such as complex biosynthetic pathways, sensors, counters and logic gates.     

Rapid loss of sensitivity to tetrodotoxin by chick ventricular myocardial cells after separation from the heart

We reported previously that chick myocardial cells placed into monolayer cell culture lost tetrodotoxin (TTX) sensitivity when tested at 72 h. To further characterize the change, ventricular myocardial cells were dispersed from chick embryos 14–16 days old; these hearts are TTX-sensitive before dispersal. Intracellular microelectrode penetrations were made into spontaneously beating cells at 9–72 h after culturing. No TTX-sensitive cells were found. Spontaneous action potentials with concomitant contractions continued in the presence of TTX (8 μg/ml), and the maximum rate of rise of the action potentials (+ _max) (control of 2–20 V/sec) was not reduced. Since the cells did not adhere to the vessel before 9 h, suspensions of cells were studied 1–8 h after dispersal to determine the rapidity of the loss of TTX sensitivity; all cells which contracted spontaneously or responded to electrical stimulation continued to beat in TTX. Addition of cycloheximide or actinomycin D did not prevent the loss of TTX sensitivity. The loss is not due to the use of trypsin (0.01 %) because dispersal by collagenase also resulted in loss of TTX sensitivity. Furthermore, cells separated mechanically (from 8-day-old hearts) also lost TTX sensitivity. In addition, loss of TTX sensitivity did not occur in frog sartorius muscles organ cultured for several days in 0.01 % trypsin. The loss of TTX sensitivity occurred even in multilayered cell cultures. Chronic exposure to carbachol or isoproterenol did not prevent the loss. However, elevation of K⁺ in the medium (12–60 mM) prevented or reversed the loss of TTX sensitivity in some cells (˜50 %), although + _max remained low. Hence, the loss of TTX-sensitive fast Na⁺ channels upon cell dispersal (a) occurs very rapidly (less than 60 min), (b) is not due to the use of trypsin, (c) is independent of protein synthesis, (d) is not solely a function of cell association, (e) is not influenced by neurotransmitters, and (f) is prevented or reversed by culturing in elevated [K⁺]₀. The mechanism of the changes in characteristics of the cation channels remains to be elucidated.     

Rapid loss of genetic variation in a founding population of Primula elatior (Primulaceae) after colonization

Background and Aims

Land-use changes and associated extinction/colonization dynamics can have a large impact on population genetic diversity of plant species. The aim of this study was to investigate genetic diversity in a founding population of the self-incompatible forest herb Primula elatior and to elucidate the processes that affect genetic diversity shortly after colonization.

Methods

AFLP markers were used to analyse genetic diversity across three age classes and spatial genetic structure within a founding population of P. elatior in a recently established stand in central Belgium. Parentage analyses were used to assess the amount of gene flow from outside the population and to investigate the contribution of mother plants to future generations.

Results

The genetic diversity of second and third generation plants was significantly reduced compared with that of first generation plants. Significant spatial genetic structure was observed. Parentage analyses showed that <20 % of the youngest individuals originated from parents outside the study population and that >50 % of first and second generation plants did not contribute to seedling recruitment.

Conclusions

These results suggest that a small effective population size and genetic drift can lead to rapid decline of genetic diversity of offspring in founding populations shortly after colonization. This multigenerational study also highlights that considerable amounts of gene flow seem to be required to counterbalance genetic drift and to sustain high levels of genetic diversity after colonization in recently established stands.Key words: AFLP, colonization, forest regeneration, genetic diversity, genetic drift, parentage analysis, spatial genetic structure     

Synthesis and carbonic anhydrase inhibitory properties of sulfamides structurally related to dopamine

A series of novel sulfamides incorporating the dopamine scaffold were synthesized. Reaction of amines and tert-butyl-alcohol/benzyl alcohol in the presence of chlorosulfonyl isocyanate (CSI) afforded sulfamoyl carbamates, which were converted to the title compounds by treatment with trifluoroacetic acid or by palladium-catalyzed hydrogenolysis. Inhibition of six α-carbonic anhydrases (CAs, EC 4.2.1.1), that is, CA I, CA II, CA VA, CA IX, CA XII and CA XIV, and two β-CAs from Candida glabrata (CgCA) and Mycobacterium tuberculosis (Rv3588) with these sulfamides was investigated. All CA isozymes were inhibited in the low micromolar to nanomolar range by the dopamine sulfamide analogues. K_is were in the range of 0.061–1.822 μM for CA I, 1.47–2.94 nM for CA II, 2.25–3.34 μM for CA VA, 0.041–0.37 μM for CA IX, 0.021–1.52 μM for CA XII, 0.007–0.219 μM for CA XIV, 0.35–5.31 μM for CgCA and 0.465–4.29 μM for Rv3588. The synthesized sulfamides may lead to inhibitors targeting medicinally relevant CA isoforms with potential applications as antiepileptic, antiobesity antitumor agents or anti-infective.     

Rapid proteomic remodeling of cardiac tissue caused by total body ionizing radiation

Accidental nuclear scenarios lead to environmental contamination of unknown level. Immediate radiation‐induced biological responses that trigger processes leading to adverse health effects decades later are not well understood. A comprehensive proteomic analysis provides a promising means to identify and quantify the initial damage after radiation exposure. Early changes in the cardiac tissue of C57BL/6 mice exposed to total body irradiation were studied, using a dose relevant to both intentional and accidental exposure (3 Gy gamma ray). Heart tissue protein lysates were analyzed 5 and 24 h after the exposure using isotope‐coded protein labeling (ICPL) and 2‐dimensional difference‐in‐gel‐electrophoresis (2‐D DIGE) proteomics approaches. The differentially expressed proteins were identified by LC‐ESI‐MS‐MS. Both techniques showed similar functional groups of proteins to be involved in the initial injury. Pathway analyses indicated that total body irradiation immediately induced biological responses such as inflammation, antioxidative defense, and reorganization of structural proteins. Mitochondrial proteins represented the protein class most sensitive to ionizing radiation. The proteins involved in the initial damage processes map to several functional categories involving cardiotoxicity. This prompts us to propose that these early changes are indicative of the processes that lead to an increased risk of cardiovascular disease after radiation exposure.     

Rapid loss of genetic variation in an endangered possum

The endangered mountain pygmy possum is the only Australian marsupial that hibernates under snow cover. Most of its alpine habitat was burnt by a rare fire in 2003, and habitat loss and disturbance have also occurred owing to ski resort development. Here we show that there has been a rapid loss of genetic variation following habitat loss associated with resort development, but no detectable loss of alleles or decrease in heterozygosity following the fire.     

Effects of R- and S-apomorphine on MPTP-induced nigro-striatal dopamine neuronal loss

In order to establish whether the antioxidant and iron-chelating activities of R-apomorphine (R-APO), a D(1)-D(2) receptor agonist, may contribute to its neuroprotective property, its S-isomer, which is not a dopamine agonist, was studied. The neuroprotective property of R- and S-APO has been studied in the MPTP model of Parkinson's disease (PD). Both S-APO (0.5-1 mg/kg, subcutaneous) and R-APO (10 mg/kg) pretreatment of C57-BL mice, protected against MPTP (24 mg/kg, intraperitoneally) induced dopamine (DA) depletion and reduction in tyrosine hydroxylase (TH) activity. However, only R-APO prevented nigro-striatal neuronal cell degeneration, as indicated by the immunohistochemistry of TH positive neurones in substantia nigra and by western analysis of striatal TH content. R-APO prevented the reduction of striatal-GSH and the increase in the ratio of GSSG over total glutathione, caused by MPTP treatment. In vitro both R-APO and S-APO inhibited monoamine oxidase A and B activity at relatively high concentrations (100 and 300 micromol/L, respectively). The elevated activity of TH induced by the two enantiomers may contribute to the maintenance of normal DA levels, suggesting that one of the targets of these molecules may involve upregulation of TH activity. It is suggested that the antioxidant and iron-chelating properties, possible monoamine oxidase inhibitory actions, together with activation of DA receptors, may participate in the mechanism of neuroprotection by APO enantiomers against MPTP.     

Rapid association of cytoskeletal remodeling proteins with the developing phagosomes of human neutrophils

Phagocytosis of opsonized particles by neutrophils involves highly localized alterations in the actin cytoskeleton that result in the formation of prominent pseudopodia and the phagocytic cup. Immunofluorescence microscopy was employed to monitor the distribution of several proteins that can regulate the cytoskeleton in human neutrophils undergoing phagocytosis of opsonized Candida albicans. The small GTPase Cdc42, its inhibitory subunit Rho-GDI, the adapter protein Nck, gamma-p21-activated protein kinase (gamma-Pak), and cofilin were found to undergo rapid association with the developing phagosomes in these cells. In contrast, these signaling proteins were either diffusely distributed in the cytoplasm or enriched in focal points at the base of the cell when optical sections were obtained from regions of the cell not involved in phagocytosis. These results are consistent with Cdc42 being critically involved in initiating the early events in phagocytosis by its ability to activate Pak and the Wiskott-Aldrich Syndrome protein that triggers the localized polymerization of actin. These data provide insights into the molecular mechanisms that trigger changes in the actin cytoskeleton during phagocytosis.