TRIP8b splice forms act in concert to regulate the localization and expression of HCN1 channels in CA1 pyramidal neurons

HCN1 channel subunits, which contribute to the hyperpolarization-activated cation current (Ih), are selectively targeted to distal apical dendrites of hippocampal CA1 pyramidal neurons. Here, we addressed the importance of the brain-specific auxiliary subunit of HCN1, TRIP8b, in regulating HCN1 expression and localization. More than ten N-terminal splice variants of TRIP8b exist in brain and exert distinct effects on HCN1 trafficking when overexpressed. We found that isoform-wide disruption of the TRIP8b/HCN1 interaction caused HCN1 to be mistargeted throughout CA1 somatodendritic compartments. In contrast, HCN1 was targeted normally to CA1 distal dendrites in a TRIP8b knockout mouse that selectively lacked exons 1b and 2. Of the two remaining hippocampal TRIP8b isoforms, TRIP8b(1a-4) promoted HCN1 surface expression in dendrites, whereas TRIP8b(1a) suppressed HCN1 misexpression in axons. Thus, proper subcellular localization of HCN1 depends on its differential additive and subtractive sculpting by two isoforms of a single auxiliary subunit.     

Dealing with scarce data to understand how environmental gradients and propagule pressure shape fine‐scale alien distribution patterns on coastal dunes

Questions: On sandy coastal habitats, factors related to substrate and to wind action vary along the sea–inland ecotone, forming a marked directional disturbance and stress gradient. Further, input of propagules of alien plant species associated to touristic exploitation and development is intense. This has contributed to establishment and spread of aliens in coastal systems. Records of alien species in databases of such heterogeneous landscapes remain scarce, posing a challenge for statistical modelling. We address this issue and attempt to shed light on the role of environmental stress/disturbance gradients and propagule pressure on invasibility of plant communities in these typical model systems. Location: Sandy coasts of Lazio (Central Italy). Methods: We proposed an innovative methodology to deal with low prevalence of alien occurrence in a data set and high cost of field‐based sampling by taking advantage, through predictive modelling, of the strong interrelation between vegetation and abiotic features in coastal dunes. We fitted generalized additive models to analyse (1) overall patterns of alien occurrence and spread and (2) specific patterns of the most common alien species recorded. Conclusion: Even in the presence of strong propagule pressure, variation in local abiotic conditions can explain differences in invasibility within a local environment, and intermediate levels of natural disturbance and stress offer the best conditions for spread of alien species. However, in our model system, propagule pressure is actually the main determinant of alien species occurrence and spread. We demonstrated that extending the information of environmental features measured in a subsample of vegetation plots through predictive modelling allows complex questions in invasion biology to be addressed without requiring disproportionate funding and sampling effort.     

Guanine nucleotide- and muscarinic agonist-dependent phosphoinositide metabolism in synaptoneurosomes from cerebral cortex of immature rats

Guanine nucleotide-, neurotransmitter-, and fluoride-stimulated accumulation of [³H]inositol phosphates ([³H]InsPs) was measured in [³H]inositol-labeled synaptoneurosomes from cerebral cortex of immature (7-day-old) and adult rats, in order to clarify the role of GTP-binding proteins (G-proteins) in modulating phosphoinositide (PtdIns) metabolism during brain development. GTP(S) [Guanosine 5-O-(3-thio)triphosphate] time- and concentration-dependently stimulated PtdIns hydrolysis. Its effect was potentiated by full (carbachol, metacholine) and partial (oxotremorine) cholinergic agonists through activation of muscarinic receptors. The presence of deoxycholate was required to demonstrate agonist protentiation of the guanine nucleotide effect. The response to GTP(S) was higher in adult than in immature rats, while the effect of cholinergic agonists was similar at the two ages examined. At both ages, histamine potentiated the effect of GTP(S), while norepinephrine was ineffective. At both ages, guanosine 5-O-(2-thio)diphosphate [GDP(S)] and pertussis toxin significantly decreased GTP(S)-induced [³H]InsPs formation. The phorbol ester phorbol 12-myristate 13-acetate (PMA), on the other hand, did not inhibit the guanine nucleotide response in synaptoneurosomes from immature rats. NaF mimicked the action of GTP(S) in stimulating PtdIns hydrolysis. Its effect was not affected by carbachol and was highly synergistic with that of AlCl₃, according to the concept that fluoroaluminate (AlF₄ ^–) is the active stimulatory species. No quantitative differences were found in the response to these salts between immature and adult animals. These results provide evidence that, in both the immature and adult rat brain, neuroreceptor activation is coupled to PtdIns hydrolysis through modulatory G-proteins.     

The cellular basis for immune interferon production in autoimmune MRL-Ipr/Ipr mice

Disturbances in immune interferon (IFN gamma) activity have been implicated in the development of human systemic lupus erythematosus (SLE) and the spontaneous disease sustained by autoimmune-prone mice. We therefore investigated the cellular basis for IFN gamma production in MRL-Ipr/Ipr mice and examined the relationship between synthesis of interleukin 2 (IL 2) and IFN gamma. In vitro IL 2 and IFN gamma production in 3 to 6-mo-old, autoimmune MRL-Ipr/Ipr and MRL-+/+ mice was compared with that seen in age- and sex-matched, immunologically normal CBA/J mice. 5 X 10(6) spleen cells were pulsed with 5 micrograms of concanavalin A (Con A), and the cellfree supernatant was assayed for IL 2 and IFN gamma activity at various times up to 72 hr. We found that peak levels of IL 2 in MRL mice were less than 10% of those in the CBA/J. Yet, production of IFN gamma by cells from the autoimmune and normal strains was quite comparable. The addition of murine IL 2 to optimally Con A-stimulated cells from the MRL-Ipr/Ipr or normal mice did not affect the subsequent peak production of IFN gamma. Although the primary producers of IFN gamma in cultures of normal mice bear the Lyt-2+ phenotype, the Lyt-1+2- T-cell subset was found to be the principal source of IFN gamma in the aged MRL-Ipr/Ipr. These data suggest that Lyt-1+ cells from MRL-Ipr/Ipr mice may be differentially responsive to the signal delivered by the same mitogenic lectin with respect to lymphokine production and may indicate a distorted commitment of such cells toward production of IFN gamma and repression of IL 2 synthesis. The relationship between hypoproduction of IL 2, this usual source of IFN gamma, and the autoimmune disease sustained by MRL-Ipr/Ipr mice remains unclear.     

Supramaximal exercise mobilizes hematopoietic progenitors and reticulocytes in athletes

Marathon runners show increased circulating CD34+ cell counts and postexercise release of interleukin-6 (IL-6), granulocyte-colony stimulating factor (G-CSF) and flt3-ligand (Bonsignore MR, Morici G, Santoro A, Pegano M, Cascio L, Bonnano A, Abate P, Mirabella F, Profita M, Insalaco G, Gioia M, Vignola AM, Majolino I, Testa U, and Hogg JC. J Appl Physiol 93: 1691-1697, 2002). In the present study we hypothesized that supramaximal ("all-out") exercise may acutely affect circulating progenitors and reticulocytes and investigated possible mechanisms involved. Progenitor release was measured by flow cytometry (n = 20) and clonogenic assays (n = 6) in 20 young competitive rowers (13 M, 7 F, age +/- SD: 17.1 +/- 2.1 yr, peak O2 consumption: 56.5 +/- 11.4 ml.min(-1).kg(-1)) at rest and shortly after 1,000 m "all-out." Release of reticulocytes, cortisol, muscle enzymes, neutrophil elastase, and several cytokines/growth factors was measured. Supramaximal exercise doubled circulating CD34+ cells (rest: 7.6 +/- 3.0, all-out: 16.3 +/- 9.1 cells/mul, P < 0.001), and increased immature reticulocyte fractions; AC133+ cells doubled, suggesting release of angiogenetic precursors. Erythrocyte burst forming units and colony forming units for granulocytes-monocytes and all blood series increased postexercise by 3.4-, 5.5-, and 4.8-fold, respectively (P < 0.01 for all). All-out rowing acutely increased plasma cortisol, neutrophil elastase, flt3-ligand, hepatocyte growth factor, VEGF, and transforming growth factor-beta1, and decreased erythropoietin; K-ligand, stromal-derived factor-1, IL-6, and G-CSF were unchanged. Therefore, all-out exercise is a physiological stimulus for progenitor release in athletes. Release of reticulocytes and proangiogenetic cells and mediators suggests tissue hypoxia as possibly involved in progenitor mobilization.     

Activation of NMDA receptors induces protein kinase A-mediated phosphorylation and degradation of matrin 3. Blocking these effects prevents NMDA-induced neuronal death

Activation of NMDA receptors leads to activation of cAMP-dependent protein kinase (PKA). The main substrates phosphorylated by PKA following NMDA receptor activation remain unidentified. The aim of this work was to identify a major substrate phosphorylated by PKA following NMDA receptor activation in cerebellar neurones in culture, and to assess whether this phosphorylation may be involved in neuronal death induced by excessive NMDA receptor activation. The main PKA substrate following NMDA receptor activation was identified by MALDI-TOFF fingerprinting as the nuclear protein, matrin 3. PKA-mediated phosphorylation of matrin 3 is followed by its degradation. NMDA receptor activation in rat brain in vivo by ammonia injection also induced PKA-mediated matrin 3 phosphorylation and degradation in brain cell nuclei. Blocking NMDA receptors in brain in vivo with MK-801 reduced basal phosphorylation of matrin 3, suggesting that it is modulated by NMDA receptors. Inhibition of PKA with H-89 prevents NMDA-induced phosphorylation and degradation of matrin 3 as well as neuronal death. These results suggest that PKA-mediated phosphorylation of matrin 3 may serve as a rapid way of transferring information from synapses containing NMDA receptors to neuronal nuclei under physiological conditions, and may contribute to neuronal death under pathological conditions.