Glial control of endocannabinoid heterosynaptic modulation in hypothalamic magnocellular neuroendocrine cells.

Cannabinoid receptors are functionally operant at both glutamate and GABA synapses on hypothalamic magnocellular neuroendocrine cells; however, retrograde endocannabinoid actions are evoked at only glutamate synapses. We tested whether the functional targeting of evoked retrograde endocannabinoid actions to glutamate, and not GABA, synapses on magnocellular neurons is the result of the spatial restriction of extracellular endocannabinoids by astrocytes. Whole-cell GABA synaptic currents were recorded in magnocellular neurons in rat hypothalamic slices following manipulations to reduce glial buffering of extracellular signals. Depolarization- and glucocorticoid-evoked retrograde endocannabinoid suppression of synaptic GABA release was not detected under normal conditions, but occurred in both oxytocin and vasopressin neurons under conditions of attenuated glial coverage and depressed glial metabolic function, suggesting an emergent endocannabinoid modulation of GABA synapses with the loss of astrocyte function. Tonic endocannabinoid suppression of GABA release was insensitive to glial manipulation. Blocking cannabinoid transport mimicked, and increasing the extracellular viscosity reversed, the effect of suppressed glial buffering on the endocannabinoid modulation of GABA release. Evoked, but not tonic, endocannabinoid modulation of GABA synapses was mediated by 2-arachidonoylglycerol. Therefore, depolarization- and glucocorticoid-evoked 2-arachidonoylglycerol release from magnocellular neurons is spatially restricted to glutamate synapses by astrocytes, but spills over onto GABA synapses under conditions of reduced astrocyte buffering; tonic endocannabinoid modulation of GABA release, in contrast, is likely mediated by anandamide and is insensitive to astrocytic buffering. Astrocytes, therefore, provide dynamic control of stimulus-evoked 2-arachidonoylglycerol, but not tonic anandamide, regulation of GABA synaptic inputs to magnocellular neuroendocrine cells under different physiological conditions.

Short-term potentiation of GABAergic synaptic inputs to vasopressin and oxytocin neurones.

The magnocellular vasopressin (VP) and oxytocin (OT) neurones undergo long-term synaptic plasticity to accommodate prolonged hormone demand. By contrast, rapidly induced,transient synaptic plasticity in response to brief stimuli could enable the activation of magnocellular neurones in response to acute challenges. Here, we report a robust short-term potentiation of asynchronous GABAergic synaptic inputs (STP(GABA)) to VP and OT neurones of the hypothalamic supraoptic nucleus elicited by repetitive extracellular electrical stimulation.The STP(GABA) required extracellular Ca2+, but did not require activation of glutamate, VP or OT receptors or nitric oxide synthesis. Presynaptic action potential generation was necessary for the induction, but not the maintenance, of STP(GABA). The STP(GABA) led to a minutes-long GABA(A)receptor-dependent increase in spike frequency in VP neurones, but not in OT neurones,consistent with an excitatory function of GABA in only VP neurones and with the generation of prolonged bursts of action potentials in VP neurones. Therefore, this short-term plasticity of GABAergic synaptic inputs is likely to play very different roles in the regulation of OT and VP neurones and their distinct patterns of physiological activation.

GABA is excitatory in adult vasopressinergic neuroendocrine cells.

Neuronal excitability in the adult brain is controlled by a balance between synaptic excitation and inhibition mediated by glutamate and GABA, respectively. While generally inhibitory in the adult brain, GABA(A) receptor activation is excitatory under certain conditions in which the GABA reversal potential is shifted positive due to intracellular Cl(-) accumulation, such as during early postnatal development and brain injury. However, the conditions under which GABA is excitatory are generally either transitory or pathological. Here, we reveal GABAergic synaptic inputs to be uniformly excitatory in vasopressin (VP)-secreting magnocellular neurons in the adult hypothalamus under normal conditions. The GABA reversal potential (E(GABA)) was positive to resting potential and spike threshold in VP neurons, but not in oxytocin (OT)-secreting neurons. The VP neurons lacked expression of the K(+)-Cl(-) cotransporter 2 (KCC2), the predominant Cl(-) exporter in the adult brain. The E(GABA) was unaffected by inhibition of KCC2 in VP neurons, but was shifted positive in OT neurons, which express KCC2. Alternatively, inhibition of the Na(+)-K(+)-Cl(-) cotransporter 1 (NKCC1), a Cl(-) importer expressed in most cell types mainly during postnatal development, caused a negative shift in E(GABA) in VP neurons, but had no effect on GABA currents in OT neurons. GABA(A) receptor blockade caused a decrease in the firing rate of VP neurons, but an increase in firing in OT neurons. Our findings demonstrate that GABA is excitatory in adult VP neurons, suggesting that the classical excitation/inhibition paradigm of synaptic glutamate and GABA control of neuronal excitability does not apply to VP neurons.

Impact of adjuvant chemotherapy in stage IB non-small-cell lung cancer: an analysis of 112 consecutively treated patients.

PURPOSE: The impact of adjuvant chemotherapy (CT) in the management of radically resected stage IB non-small cell lung cancer (NSCLC) is highly debated. The aim of this study was to evaluate the outcome of this category of patients treated at our institution. METHODS: We retrospectively analysed the survival data of patients with pathologic stage IB NSCLC, who received at least 1 cycle of adjuvant CT. CT was planned to be platinum based and to be delivered for 6 cycles. RESULTS: One hundred and twelve consecutively treated patients were evaluated. PATIENT CHARACTERISTICS: median age 60 years, median tumor diameter 4 cm, 87% underwent lobectomy and 13% pneumonectomy, 58% had visceral pleural involvement (VPI). After a median follow up of 46 months, the estimated 5-year disease-free (DFS) and overall survival (OS) rates were 68% and 77%, respectively. The mean number of CT cycles was 5.2 (range 3-6), with 82% of patients receiving ≥ 5 cycles. The median cisplatin dose intensity (DI) was 22 mg/m(2)/week, and the relative DI was 85%. Median total cisplatin (CDDP) dose/patient was 416 mg/m(2). A total of 31 (27.6%) relapses were recorded, of which 81% were distant. Multivariate analysis showed no significant interaction between overall survival and the following variables: gender, type of surgery, histology, tumor volume, VPI. CONCLUSION: Our results compare favorably with the historical data evaluating the outcome of stage IB patients treated by surgery alone in a customary medical setting. Overall, our data support the use of adjuvant CT in stage IB NSCLC patients.

Oxidative photodamage induced by photodynamic therapy with methoxyphenyl porphyrin derivatives in tumour-bearing rats.

The oxidative effects of photodynamic therapy with 5,10,15,20-tetrakis(4-methoxyphenyl) porphyrin (TMP) and Zn-5,10,15,20-tetrakis(4-methoxyphenyl) porphyrin (ZnTMP) were evaluated in Wistar rats subcutaneously inoculated with Walker 256 carcinoma. The animals were irradiated with red light (λ = 685 nm; D = 50 J/cm2; 15 min) 3 h after intra-peritoneal administration of 10 mg/kg body weight of porphyrins. The presence of free radicals in tumours after photodynamic therapy with TMP and ZnTMP revealed by chemiluminescence of luminol attained the highest level at 18 h after irradiation. Lipid peroxides measured as thiobarbituric-reactive substances and protein carbonyls, which are indices of oxidative effects produced on susceptible biomolecules, were significantly increased in tumour tissues of animals 24 h after photodynamic therapy. The levels of thiol groups and total antioxidant capacity in the tumours were decreased. The activities of antioxidant enzymes superoxide dismutase and glutathione peroxidase were also increased in tumour tissues after photodynamic therapy. Increased levels of plasma lipid peroxides as well as changes in the levels of erythrocyte antioxidant enzyme activities suggest possible systemic effects of photodynamic therapy with TMP and ZnTMP.

Increased intrinsic excitability and decreased synaptic inhibition in aged somatosensory cortex pyramidal neurons.

Sensorimotor performance declines during advanced age, partially due to deficits in somatosensory acuity. Cortical receptive field expansion contributes to somatosensory deficits, suggesting increased excitability or decreased inhibition in primary somatosensory cortex (S1) pyramidal neurons. To ascertain changes in excitability and inhibition, we measured both properties in neurons from vibrissal S1 in brain slices from young and aged mice. Because adapting and non-adapting neurons-the principal pyramidal types in layer 5 (L5)-differ in intrinsic properties and inhibitory inputs, we determined age-dependent changes according to neuron type. We found an age-dependent increase in intrinsic excitability in adapting neurons, caused by a decrease in action potential threshold. Surprisingly, in non-adapting neurons we found both an increase in excitability caused by increased input resistance, and a decrease in synaptic inhibition. Spike frequency adaptation, already small in non-adapting neurons, was further reduced by aging, whereas sag, a manifestation of Ih, was increased. Therefore, aging caused both decreased inhibition and increased intrinsic excitability, but these effects were specific to pyramidal neuron type.

Synchronized bursts of miniature inhibitory postsynaptic currents.

Spike-independent miniature postsynaptic currents are generally stochastic and are therefore not thought to mediate information relay in neuronal circuits. However, we recorded endogenous bursts of IPSCs in hypothalamic magnocellular neurones in the presence of TTX, which implicated a coordinated mechanism of spike-independent GABA release. IPSC bursts were identical in the absence of TTX, although the burst incidence increased 5-fold, indicating that IPSC bursts were composed of miniature IPSCs (mIPSCs), and that the probability of burst generation increased with action potential activity. IPSC bursts required extracellular calcium, although they were not dependent on calcium influx through voltage-gated calcium channels or on calcium mobilization from intracellular stores. Current injections simulating IPSC bursts were capable of triggering and terminating action potential trains. In 25% of dual recordings, a subset of IPSC bursts were highly synchronized in onset in pairs of magnocellular neurones. Synchronized IPSC bursts displayed properties that were consistent with simultaneous release at GABA synapses shared between pairs of postsynaptic magnocellular neurones. Synchronized bursts of inhibitory synaptic inputs represent a novel mechanism that may contribute to the action potential burst generation, termination and synchronization responsible for pulsatile hormone release from neuroendocrine cells.

In vitro biocompatibility and corrosion resistance of a new implant titanium base alloy.

One objective of this work was to study the corrosion resistance of the new implant Ti-10Zr-5Ta-5Nb alloy in physiological fluids of different pH values, simulating the extreme functional conditions. Another objective was in vitro biocompatibility evaluation of the new alloy using human fetal osteoblast cell line hFOB 1.19. Cytocompatibility was assessed by determination of possible material cytotoxic effects, cell morphology and cell adhesion. The thermo-mechanical processing of the new implant alloy consisted in plastic deformation (almost 90%) performed by hot rolling accompanied by an initial and final heat treatment. The new Ti-10Zr-5Ta-5Nb alloy presented self-passivation, with a large passive potential range and low passive current densities, namely, a very good anticorrosive resistance in Ringer solution of acid, neutral and alkaline pH values. Cell viability was not affected by the alloy substrate presence and a very good compatibility was noticed.

Controversies around the use of monoclonal antibodies in the treatment of advanced non-small cell lung cancer.

First line combination chemotherapy (CT) using platinum-based doublets is established as a standard of care for advanced non-small cell lung cancer (NSCLC). Nevertheless, no significant advances have been recorded during the last years in this field. Therefore, there is a wide consensus among physicians that a plateau has already been reached with this strategy. Targeted therapy using tyrosine-kinase inhibitors (TKIs) and monoclonal antibodies emerged as a new field of development in the NSCLC therapeutics. Recently, the results of the phase III trials testing antibodies against vascular endothelial growth factor-VEGF (bevacizumab) and epidermal growth factor receptor-EGFR (cetuximab) challenged the paradigm of the platinum doublets as a gold standard in advanced NSCLC. Their appearance was enthusiastically commended both by patients and the oncological community. However, all medical oncologists have the responsibility to carefully analyze the real benefits of these new agents, to balance the advantages against the implicit risks of therapy and to make the decision having in mind the best interest of their patients. Last but not least, the associated health economic burden should also be considered. This paper addresses some issues related to the use of cetuximab and bevacizumab in advanced NSCLC. The main controversial aspects regarding patient selection, the real benefit of therapy, the molecular and clinical predictors, and the impact of other independent variables are carefully examined and presented. Due to many unsolved questions, no definite conclusions can be supported. The final decision about the optimal use of these agents is left to the clinical judgment of each treating physician.

Lactation induces increased IPSC bursting in oxytocinergic neurons.

Hypothalamic magnocellular neurosecretory cells (MNCs) undergo dramatic structural reorganization during lactation in female rats that is thought to contribute to the pulsatile secretion of oxytocin critical for milk ejection. MNCs from male rats generate robust bursts of GABAergic synaptic currents, a subset of which are onset-synchronized between MNC pairs, but the functional role of the IPSC bursts is not known. To determine the physiological relevance of IPSC bursts, we compared MNCs from lactating and non-lactating female rats using whole-cell recordings in brain slices. We recorded a sixfold increase in the incidence of IPSC bursts in oxytocin (OT)-MNCs from lactating rats compared to non-lactating rats, whereas there was no change in IPSC bursts in vasopressin (VP)-MNCs. Synchronized bursts of IPSCs were observed in pairs of MNCs in slices from lactating rats. Our data indicate, therefore, that IPSC bursts are upregulated specifically in OT-MNCs during lactation, and may, therefore, contribute via rebound depolarization to the spike trains in OT neurons that lead to reflex milk ejection.

An investigation on performance of Significance Analysis of Microarray (SAM) for the comparisons of several treatments with one control in the presence of small-variance genes.

One of multiple testing problems in drug finding experiments is the comparison of several treatments with one control. In this paper we discuss a particular situation of such an experiment, i.e., a microarray setting, where the many-to-one comparisons need to be addressed for thousands of genes simultaneously. For a gene-specific analysis, Dunnett's single step procedure is considered within gene tests, while the FDR controlling procedures such as Significance Analysis of Microarrays (SAM) and Benjamini and Hochberg (BH) False Discovery Rate (FDR) adjustment are applied to control the error rate across genes. The method is applied to a microarray experiment with four treatment groups (three microarrays in each group) and 16,998 genes. Simulation studies are conducted to investigate the performance of the SAM method and the BH-FDR procedure with regard to controlling the FDR, and to investigate the effect of small-variance genes on the FDR in the SAM procedure.

Publisher Correction: Marked bias towards spontaneous synaptic inhibition distinguishes non-adapting from adapting layer 5 pyramidal neurons in the barrel cortex.

A correction to this article has been published and is linked from the HTML version of this paper. The error has been fixed in the paper.

In Vitro Cytotoxicity of Zinc Fructoborate, a Novel Zinc-Boron Active Natural Complex.

In recent years, the role of zinc in biological systems has been a subject of intense research. Zinc is required for multiple metabolic processes as a structural, regulatory, or catalytic ion. The objective of this study, was to assess the toxicity profile of a newly synthesized zinc-boron molecule on cultured cells. Zinc fructoborate, at different levels of concentration, was tested for its impact on the Vero kidney cell line (ATCC® CCL-81™) using the MTT assay. The compound exhibited a low cytotoxic effect on the cell line. Thus, our study demonstrates that the zinc fructoborate could become a promising dietary supplement molecule.

Marked bias towards spontaneous synaptic inhibition distinguishes non-adapting from adapting layer 5 pyramidal neurons in the barrel cortex.

Pyramidal neuron subtypes differ in intrinsic electrophysiology properties and dendritic morphology. However, do different pyramidal neuron subtypes also receive synaptic inputs that are dissimilar in frequency and in excitation/inhibition balance? Unsupervised clustering of three intrinsic parameters that vary by cell subtype - the slow afterhyperpolarization, the sag, and the spike frequency adaptation - split layer 5 barrel cortex pyramidal neurons into two clusters: one of adapting cells and one of non-adapting cells, corresponding to previously described thin- and thick-tufted pyramidal neurons, respectively. Non-adapting neurons presented frequencies of spontaneous inhibitory postsynaptic currents (sIPSCs) and spontaneous excitatory postsynaptic currents (sEPSCs) three- and two-fold higher, respectively, than those of adapting neurons. The IPSC difference between pyramidal subtypes was activity independent. A subset of neurons were thy1-GFP positive, presented characteristics of non-adapting pyramidal neurons, and also had higher IPSC and EPSC frequencies than adapting neurons. The sEPSC/sIPSC frequency ratio was higher in adapting than in non-adapting cells, suggesting a higher excitatory drive in adapting neurons. Therefore, our study on spontaneous synaptic inputs suggests a different extent of synaptic information processing in adapting and non-adapting barrel cortex neurons, and that eventual deficits in inhibition may have differential effects on the excitation/inhibition balance in adapting and non-adapting neurons.

Highly dissimilar behaviors mediated by a multifunctional network in the marine mollusk Tritonia diomedea.

Several motor networks have now been found to be multifunctional, in which one group of neurons participates in the generation of multiple behavioral motor programs. Not surprisingly, the behaviors involved are frequently closely related, often using the same or similar muscle groups. Here we describe an interneuronal network in the marine mollusk Tritonia diomedea that is involved in producing two highly dissimilar behaviors, rhythmic, muscle-based escape swimming and nonrhythmic, cilia-mediated crawling. Several observations support this conclusion. First, the dorsal swim interneurons (DSIs) of the swim central pattern generator (CPG) directly excite Pedal neuron 21 (Pd21) and Pd5, the only identified cilia-activating efferent neurons in Tritonia. Second, stimulation of a single DSI elicits beating of the foot cilia in semi-intact preparations and crawling in intact animal treadmill preparations. Third, the DSIs fire at an elevated rate for nearly 1 hr after a swim motor program, which correlates reasonably well with the period freely behaving animals were found to crawl after they swam. Fourth, silencing the tonically active DSIs after a swim motor program substantially reduces or eliminates ongoing cilia neuron firing, indicating that the DSIs are major contributors to the synaptic input driving these cells. Finally, all of the other swim CPG neurons also connect to the cilia neurons, most monosynaptically. Taken together, these observations indicate that the Tritonia swim CPG network participates in producing both escape swimming and crawling. Given the extreme differences between these behaviors---rhythmic versus tonic, muscular versus ciliary, and brief versus prolonged--these findings reveal a striking versatility for a small multifunctional network.

Factors influencing student nurse decisions to report poor practice witnessed while on placement.

BACKGROUND: While it is commonly accepted that nursing care is generally of a good standard, it would be naïve to think that this is always the case. Over recent years, concern about aspects of the quality of some nursing care has grown. In tandem with this, there is recognition that nurses do not always report poor practice. As future registrants, student nurses have a role to play in changing this culture. We know, however, relatively little about the factors that influence student decisions on whether or not to report. In the absence of a more nuanced understanding of this issue, we run the risk of assuming students will speak out simply because we say they should. OBJECTIVES: To explore influences on student decisions about whether or not to report poor clinical practice, which is a result of deliberate action and which is witnessed while on placement. METHODS: Qualitative interviews were conducted with thirteen pre-registration nursing students from the UK. Participants included both adult and mental health nurses with an age range from 20 to 47. Data were analysed to identify key themes. Category integrity and fit with data were confirmed by a team member following initial analysis. RESULTS: Four themes. The first of these, 'I had no choice' described the personal and ethical drivers which influenced students to report. 'Consequences for self' and 'Living with ambiguity' provide an account of why some students struggle to report, while 'Being prepared' summarised arguments both for and against reporting concerns. CONCLUSION: While there is a drive to promote openness in health care settings and an expectation that staff will raise concerns the reality is that the decision to do this can be very difficult. This is the case for some student nurses. Our results suggest ways in which educationalists might intervene to support students who witness poor practice to report.

Spectral analysis of the porphyrins incorporation into human blood.

The porphyrins seem to be the most effective photosensitizers in photodynamic therapy of cancer. Monomers and dimers of sulfonated and nonsulfonated porphyrins [5,10,15,20-tetra-phenyl-porphyrin, 5,10,15,20-tetra-naphtyl-porphyrin (TNP), 5,10,15,20-tetra-p-sulfonato-phenyl-porphyrin (TS4PP) and 5,10,15,20-tetra-p-sulfonato-naphtyl-porphyrin], are studied in this paper by means of different spectral methods (UV-vis, fluorescence and polarization fluorescence). The porphyrins as TNP seem to be the most effective photosensitizer especially in a DMSO:water binary mixture of solvent. Monomer-dimer and J-aggregation equilibria and the temperature dependence of TS4PP spectra are also studied in this paper. The incorporation of the forms of these porphyrins into blood cells is studied by means of the cytofluorimetric method. © 1999 Society of Photo-Optical Instrumentation Engineers.

The incorporation of various porphyrins into blood cells measured via flow cytometry, absorption and emission spectroscopy.

The incorporation of the five following porphyrins: meso-tetra(4-phenyl)porphyrin (TPP); meso-tetra(4-sulfonato-phenyl)porphyrin (TPPS4); meso-tetra(4-naphthyl)porphyrin (TNP); tri-sulfo-tetra-phenyl porphyrin (TPPS3) and tetra-sulfonato-naphthyl porphyrin (TNPS4) into human blood cells was investigated using flow cytometry, and absorption and emission spectroscopy. The percentage of stained cells, measured in a fluorescence cytometer, provided information on the efficiency of incorporation of fluorescent dye molecules into different types of cells. The yield of the incorporation of a dye was dependent on the type of dye and the solvent used for cell incubation. The degree of dye aggregation and ionization varied with the incubation medium, but dye molecules incorporated into cells seemed to be restricted to those in the monomeric state, exhibiting similar fluorescence yield. Of the three sulfonated porphyrins investigated only TPPS4 was efficiently incorporated into leukocytes. In the incubation solvent, this dye was in monomeric and neutral form. TPPS3 which was also in monomeric form, practically was not incorporated into cells. TPP and TNP dissolved in 5% aqueous dimethyl sulfoxide were present mostly in aggregated forms but they penetrated the cells with a high efficiency.

The interactions of phthalocyanines with stimulated and resting human peripheral blood mononuclear cells.

The interactions of two metal-free phthalocyanines [(H2Pc) and Solar Pc (with four peripherical groups: SO2N(CH2CH2OH)2)] and of one metal substituted dye (CoPc) with resting and stimulated human peripheral blood mononuclear cells (PBMC) were compared. The absorption, fluorescence, photoacoustic and EPR spectra of both resting cells and cells stimulated by phytohaemagglutinin, incubated in dimethyl sulfoxide (DMSO) with very low or 95% water content and with or without dye addition, were measured. The fate of the light absorbed by the samples was investigated. It is known that singlet oxygen production is crucial for photodynamic action of dyes. Thermal deactivation and luminescence emission compete with this process, so investigation of these alternative paths of sensitizer deactivation provides information about photodynamic action. The incorporation of the investigated dyes into cells and the perturbation of the cell structure caused by the dyes, the incubation solvent and the activator were investigated by comparing the spectral properties of PBMC before and after stimulation and incubation. Incubation of the cells for 1 h in a solution of Solar Pc in 99.5% aqueous DMSO, resulted in an efficient dye incorporation which was highly selective. Solar Pc being introduced much more efficiently into stimulated cells than into resting cells.

The British reaction to dementia praecox 1893-1913. Part 1.

Emil Kraepelin introduced the concept of dementia praecox in 1893. The eventual acceptance of the concept brought a degree of clarity and order previously unknown to psychiatric nosology. The pre-Kraepelin era had been dominated by concepts such as mania, melancholia and adolescent insanity. After Kraepelin these ideas were abandoned in favour of the two great concepts of dementia praecox and manic depressive insanity, both of which remain active within modern psychiatry in the form of schizophrenia and bipolar disorder. This two-part study focuses on the early British reaction to Kraepelin's concept, from 1893, when he first introduced it, to 1913 when it gained general recognition. It examines the struggle experienced by the proponents of dementia praecox before the concept's acceptance by most British psychiatrists in 1913. It argues that both clinical/professional and linguistic factors influenced the British response to dementia praecox. Part 1 of this study describes the backdrop to the development of Kraepelin's ideas and examines the response to the concept in the British psychiatric textbooks and journals of the period. Part 2 will explore reaction to the concept in the professional meetings of the period, and will also examine and evaluate the key issues arising from the debate.