Differential regulation of STP, LTP and LTD by structurally diverse NMDA receptor subunit-specific positive allosteric modulators.

Different types of memory are thought to rely on different types of synaptic plasticity, many of which depend on the activation of the N-Methyl-D Aspartate (NMDA) subtype of glutamate receptors. Accordingly, there is considerable interest in the possibility of using positive allosteric modulators (PAMs) of NMDA receptors (NMDARs) as cognitive enhancers. Here we firstly review the evidence that NMDA receptor-dependent forms of synaptic plasticity: short-term potentiation (STP), long-term potentiation (LTP) and long-term depression (LTD) can be pharmacologically differentiated by using NMDAR ligands. These observations suggest that PAMs of NMDAR function, depending on their subtype selectivity, might differentially regulate STP, LTP and LTD. To test this hypothesis, we secondly performed experiments in rodent hippocampal slices with UBP714 (a GluN2A/2B preferring PAM), CIQ (a GluN2C/D selective PAM) and UBP709 (a pan-PAM that potentiates all GluN2 subunits). We report here, for the first time, that: (i) UBP714 potentiates sub-maximal LTP and reduces LTD; (ii) CIQ potentiates STP without affecting LTP; (iii) UBP709 enhances LTD and decreases LTP. We conclude that PAMs can differentially regulate distinct forms of NMDAR-dependent synaptic plasticity due to their subtype selectivity.

Towards an ASIC-based Fluoroscopic Capsule for the Early Cancer Detection in the Small Intestine.

Early micro-cancer detection in the small intestine can be realized using infrared fluorescence endoscopy (IRFE) in conjunction with an infrared fluorescence biomarker. In this paper, we present a third-generation capsule that detects weak fluorescence signals emitted by low concentrations of indocyanine green (ICG). An applicationspecific integrated circuit (ASIC) has been designed and fabricated that integrates many of the peripheral components of the capsule system. The ASIC enables the system to have greater sensitivity whilst reducing the capsule size and lowering the power consumption.

Spatial Analytic Phase Difference of EEG activity during anesthetic-induced unconsciousness.

OBJECTIVE: A number of studies provide supporting evidence for changes in synchronization during anesthetic-induced unconsciousness. This study investigates how anesthetic administration affects the widespread patterns of phase synchrony. METHODS: The recently introduced method of Spatial Analytic Phase Difference (SAPD) was used to measure changes in synchrony in the electroencephalogram (EEG) activity of 29 patients undergoing routine surgery. Analysis was performed over 9 frequency bands: (i) δ (1.5-3.5Hz); (ii) θ (3.5-7.5Hz); (iii) α1 (8-10Hz); (iv) α2 (10.5-12Hz); (v) ß1 (12.5-18Hz); (vi) ß2 (18.5-21Hz); (vii) ß3 (21.5-30Hz); (viii) γ1 (30.5-40Hz); and (ix) γ2 (60-80Hz). RESULTS: Anesthesia was characterized by (a) large and localized synchrony increases in mid-frequency bands (8-12Hz), (b) smaller and widespread synchrony increases in higher frequency bands (30.5-40Hz, 60-80Hz), and (c) both increase and decrease of synchrony in low frequency bands (1.5-7.5Hz). CONCLUSIONS: This study supports anesthetic-induced changes in synchrony, with the inducement of persistent and reversible widespread γ synchrony being most prominent. SIGNIFICANCE: Our findings have implications in the study of consciousness, support existing literature in the field and contribute towards the theoretical understanding of the mechanisms behind loss of consciousness. Future investigations could result in a synchrony-based measure for monitoring the level of hypnosis of patients during surgery.

Global field synchrony during general anaesthesia.

BACKGROUND: Anaesthetic agents may disrupt consciousness by inhibiting long-range synchronization of brain activity. In the current study, the patterns of widespread and spatially localized synchrony during anaesthesia are investigated using a measure called global field synchrony (GFS). METHODS: The EEG obtained during routine surgery in 29 patients was analysed with GFS over the following frequency bands: δ (1.5-3.5 Hz), θ (3.5-7.5 Hz), α1 (8-10 Hz), α2 (10.5-12 Hz), ß1 (12.5-18 Hz), ß2 (18.5-21 Hz), ß3 (21.5-30 Hz), γ1 (30.5-40 Hz), and γ2 (60-80 Hz). In addition, localized GFS estimations over aggregate brain areas were performed. GFS was estimated over 2 s non-overlapping windows. The differences in GFS values between 'wakefulness' and 'anaesthesia' were assessed with the two-sided Wilcoxon rank-sum tests (α=0.05). RESULTS: Anaesthetic administration caused significant GFS changes in all frequency ranges and electrode combinations studied: (i) widespread synchrony increased in the α2 and ß1 ranges and decreased in all other ranges, with the exception of α1 and ß2, where no specific pattern was identified; and (ii) localized synchrony decreased in all areas in the δ and γ2 ranges, while location-specific changes were observed in the remaining frequency ranges. The most consistent findings were statistically significant decreases over all areas in the γ2 range, with GFS decrease over the central-right temporal being the most consistent change. CONCLUSIONS: Significant frequency- and location-dependent changes in GFS were induced by anaesthetic administration, with more robust changes identified in the γ range. GFS can act as an aid for further and more detailed analysis regarding the particular combinations of frequency ranges and spatial locations that are most informative for the study of anaesthetic-induced unconsciousness.

The use of permutation entropy to characterize sleep electroencephalograms.

This work proposes the use of Permutation Entropy (PE), a measure of time-series complexity, to characterize electroencephalogram (EEG) signals recorded during sleep. Such a measure could provide information concerning the different sleep stages and, thus, be utilized as an additional aid to obtain sleep staging information. PE has been estimated for artifact-free 30s segments from more than 80 hours of EEG records obtained from 16 subjects during all-night recordings, from which the mean PE for each sleep stage was obtained. It was found that different sleep stages are characterized by significantly different PE values, which track the physiological changes in the complexity of the EEG signals observed at the different sleep stages. This finding encourages the use of PE as an additional aide to either visual or automated sleep staging.

Entropy measures for discrimination of 'awake' Vs 'anaesthetized' state in recovery from general anesthesia.

Approximate Entropy (ApEn) and Permutation Entropy (PE) have been recently introduced for assessment of anesthetic depth. Both measures have previously been shown to track changes in the electrical brain activity related to the administration of anesthetic agents. In this paper ApEn and PE are compared for the automatic classification of 'awake' and 'anesthetized' state using a Support Vector Machine to assess their robustness for potential use in a device for monitoring awareness during general anesthesia. It was found that both measures provide linearly separable features and we are able to discriminate between the two states with accuracy greater than 96% using either of the two entropy measures.

A micropower tilt-processing circuit.

This paper describes a novel analog circuit for extracting the tilt angle from the output of a linear microelectromechanical-system accelerometer. The circuit uses the accelerometer signal, together with the gravitational acceleration vector, to generate the tilt signal. Using a current-mode representation with metal-oxide semiconductor devices operating in weak inversion, the appropriate trigonometric function has been realized to compute tilt. Furthermore, implementing a long-time constant filter to extract the mean tilt level provides adaptation to the static tilt level. Specifically, this circuit has been designed as part of an implantable vestibular prosthesis to provide inclination signals for bypassing dysfunctional otolith end organs. The circuit has been fabricated in AustriaMicroSystems 0.35-mum 2P4M complementary metal-oxide semiconductor technology, and this paper presents the theory, implementation, and measured results.

A partial-current-steering biphasic stimulation driver for vestibular prostheses.

This paper describes a novel partial-current-steering stimulation circuit for implantable vestibular prostheses. The drive hardware momentarily delivers a charge-balanced asymmetric stimulus to a dummy load before steering towards the stimulation electrodes. In this fashion, power is conserved while still gaining from the benefits of current steering. The circuit has been designed to be digitally programmable as part of an implantable vestibular prosthesis. The hardware has been implemented in AMS 0.35 mum 2P4M CMOS technology.

Germ cell transplantation: a review and progress report on ICSI from spermatozoa generated in xenogeneic testes.

Results from the transplantation of donor male germ cells into xenogeneic recipient seminiferous tubules indicate that donor spermatogonia are capable of differentiating to form spermatozoa morphologically characteristic of the donor species. Germ cell transplantation procedures combined with developments in freezing, culturing or enriching germ cell populations have applications of paramount importance in medicine, basic sciences and animal reproduction. Additionally, these techniques can serve as an alternative approach for gonadal protection and fertility preservation in patients with cancer. This article is a chronological critical review of the technological advances that followed the initial successful transplantation of mouse germ cells into recipient mice. Furthermore, the factors responsible for the immunological privilege properties of the testis and the parameters influencing the potential of mammalian germ cells to undergo mitosis and meiosis within a xenogeneic testis are described. Finally, the role of human germ cell transplantation procedures in the therapeutic management of non-obstructive azoospermia is discussed.

Use of a highly sensitive quantitative telomerase assay in intracytoplasmic sperm injection programmes for the treatment of 47,XXY non-mosaic Klinefelter men.

We evaluated the role of the sensitive quantitative telomerase assay (SQTA) in the management of men with non-mosaic Klinefelter's syndrome (KS). Diagnostic testicular biopsy (DTB) was performed in 24 men with KS. A part of the DTB was stained and the remaining fragment was processed for the SQTA. After 3-18 months, a therapeutic testicular biopsy (TTB) was performed in the same testicle and the recovered specimens were processed to identify spermatozoa. Men with a SQTA outcome equal to 0.00 Units microg-1 protein (n = 7) demonstrated therapeutic testicular biopsy material that was negative for spermatogenic cells. In five men with a SQTA outcome of 8.11-38.03 Units microg-1, the most advanced germ cell was the spermatogonium/primary spermatocyte. In the remaining 12 men, the most advanced spermatogenic cell in the TTB was the spermatozoon. In these men, the SQTA outcome was equal to 25.76-92.68 Units microg-1 protein. Using 39.00 Units microg-1 protein as a cut-off value, the accuracy of the SQTA in identifying men positive for spermatozoa was 91.6%. It appears that the SQTA has a role for identifying non-mosaic KS men who have testicular spermatozoa.

The receptor tyrosine kinase EphB2 regulates NMDA-dependent synaptic function.

Members of the Eph family of receptor tyrosine kinases control many aspects of cellular interactions during development, including axon guidance. Here, we demonstrate that EphB2 also regulates postnatal synaptic function in the mammalian CNS. Mice lacking the EphB2 intracellular kinase domain showed wild-type levels of LTP, whereas mice lacking the entire EphB2 receptor had reduced LTP at hippocampal CA1 and dentate gyrus synapses. Synaptic NMDA-mediated current was reduced in dentate granule neurons in EphB2 null mice, as was synaptically localized NR1 as revealed by immunogold localization. Finally, we show that EphB2 is upregulated in hippocampal pyramidal neurons in vitro and in vivo by stimuli known to induce changes in synaptic structure. Together, these data demonstrate that EphB2 plays an important role in regulating synaptic function.

Non-myelin-forming perisynaptic schwann cells express protein zero and myelin-associated glycoprotein.

Perisynaptic Schwann cells (PSCs) envelop axonal terminals and are physiologically distinct from the nearby myelinating Schwann cells (MSCs), which surround the same innervating motor axons. PSCs have special functions at the neuromuscular synapse, where they detect and can modulate neurotransmitter release. Although PSCs are similar to non-myelinating Schwann cells in that they do not form multiple myelin wrappings around nerve terminals, they do wrap around single nerve terminals. These differences, as well as others, lead us to question whether PSCs are truly of Schwann cell origin. We thus characterized the expression of molecules, classically associated with myelin and Schwann cells, in PSCs at the frog neuromuscular junction. We wondered whether PSCs express the Schwann cell marker protein zero (P(0)) and whether their lack of myelination was related to an absence of myelin-associated glycoprotein (MAG), a protein found in myelinating cells that is considered important in myelination. Instead, we found that PSCs express both P(0) and MAG, and other myelinating glial markers such as galactocerebroside and 2',3'-cyclic nucleotide 3'-phosphodiesterase. In denervated preparations, P(0) and MAG expression persisted, including at newly formed PSC extensions. Because PSCs do not myelinate, it is clear that expression of these proteins alone is not sufficient for myelin formation. It is possible that factors present at synapses may prevent myelination, while P(0) and MAG may mediate adhesion between nerve terminals and the surrounding PSCs. The results indicate that PSCs are of Schwann cell origin.

Muscarinic control of cytoskeleton in perisynaptic glia.

Similar to astrocytes at CNS synapses, perisynaptic Schwann cells (PSCs) surround nerve terminals at the neuromuscular junction (NMJ). These special teloglial cells are sensitive to neurotransmitters and upregulate glial fibrillary acidic protein (GFAP) when deprived of synaptic activity. We found that activation of muscarinic acetylcholine receptors (mAChRs) at PSCs, but not purinergic (ATP and adenosine) or peptidergic [substance P (SP) and calcitonin gene-related peptide (CGRP)] receptors, prevented this upregulation. When applied onto single PSCs, muscarine evoked Ca2+ responses that fatigued but prevented upregulation of this glial cytoskeletal protein. Application of ATP onto single PSCs evoked Ca2+ signals that showed little fatigue, and GFAP upregulation occurred. Thus, Ca2+ signals alone cannot prevent GFAP upregulation in the PSCs. After blockade of cholinergic receptors by gallamine, neuronal activity was not effective in maintaining low GFAP levels in the perisynaptic glia. Last, immunohistochemistry disclosed mAChRs on PSCs and nearby fibroblasts. Thus, acetylcholine secreted by the nerve terminal acts on the PSCs via mAChRs to regulate GFAP. Cytoskeletal changes may influence perisynaptic glial functions, including growth, remodeling, and modulation of the synapse.

In vitro and in vivo evaluation of insulin-producing beta TC6-F7 cells in microcapsules.

In the present study, the insulin secretory capacity of beta TC6-F7 cells in microcapsules was evaluated. The cell mass within capsules was found to expand in a three-dimensional fashion, in contrast to cells seeded on plates that grew as a monolayer. In in vitro studies, both free and encapsulated cells were found to secrete insulin in the absence of glucose, at 13.6 +/- 1.1 and 14.5 +/- 0.9 ng.10(6) cells-1.60 min-1, respectively, with the response rising to a maximum of 26.0 +/- 0.8 and 31 +/- 2.3 ng.10(6) cells-1.60 min-1 in the presence of 16.8 mM glucose. Encapsulated cells were able to produce Ca2+ responses in the presence of KCl (50 mM) and BAY K 8644 (100 microM). In in vivo studies, intraperitoneal transplantation of 3.0 x 10(6) microencapsulated cells into mice (n = 5) with streptozotocin-induced diabetes resulted in the restoration of normoglycemia up to 57 days. Insulin concentrations rose from 0.4 +/- 0.1 ng/ml before the graft administration to 2.2 +/- 0.8 ng/ml after the transplantation in the normoglycemic recipients. An oral glucose challenge in transplant recipients demonstrated a flat glucose response, suggesting extremely high glucose clearance rates. These data demonstrate the potential use of the immunoisolated beta-cell lines for the treatment of diabetes.

Administration of high doses of recombinant human erythropoietin to patients with beta-thalassemia intermedia: a preliminary trial.

Four patients (1 male, 3 female, age range 16-56 yr) with beta-thalassemia intermedia were given high doses of recombinant human erythropoietin (rHuEpo), iron sulfate and folic acid in an attempt to improve their anemia. The dose schedule was: rHuEpo, 500 U/kg 3 times weekly, iron sulfate, 300 mg/d and folic acid, 5 mg/d. All patients were red blood cell transfusion-dependent. Hematological data and fetal hemoglobin (HbF) were assayed every 2 wk. XmnI polymorphism and beta-thalassemia mutations were identified by PCR. All patients showed a moderate to high increase in hemoglobin values (mean value: 2.5 g/dl) and in 1 patient HbF levels also increased; 3 patients became red blood cell transfusion-independent and 1 patient was able to extend the intervals between transfusions significantly. No side effects were observed during rHuEpo therapy.

Imaging of calcium in Drosophila larval motor nerve terminals.

Calcium measurements in the presynaptic terminal are essential in the investigation of mechanisms underlying neurotransmitter release. To enhance the genetic analysis of secretory mechanisms, we have developed Ca2+ imaging techniques for Drosophila larval motor nerve terminals. We studied Ca2+ signals in "big" (type Ib) and "small" (type Is) boutons that innervate ventral longitudinal muscles 6 and 7 in each abdominal segment of Canton-S (CS)-strain 3rd instar larvae. The indicator fluo-3 in conjunction with confocal microscopy was used to detect stimulus-dependent changes in [Ca2+]i. The Ca2+ signals were reliable and reproducible, and the resting fluorescence remained constant throughout the experiments. The Ca2+ signals increased with stimulus frequency from 5 to 20 Hz for both bouton types. No significant differences in the Ca2+ signals were seen between the two bouton types at 5 and 20 Hz, but there was a difference at 10 Hz. The decay of the Ca2+ signal was more prolonged after 20-Hz stimulation than after 5 and 10 Hz. At the single-synapse level, the secretory efficacy of Is synapses is greater than that of Ib synapses, but our data show that factors other than differences in Ca2+ entry may govern the strength of synaptic transmission.

Strength of synaptic transmission at neuromuscular junctions of crustaceans and insects in relation to calcium entry.

Crustacean and insect neuromuscular junctions typically include numerous small synapses, each of which usually contains one or more active zones, which possess voltage-sensitive calcium channels and are specialized for release of synaptic vesicles. Strength of transmission (the number of quantal units released per synapse by a nerve impulse) varies greatly among different endings of individual neurons, and from one neuron to another. Ultrastructural features of synapses account for some of the physiological differences at endings of individual neurons. The nerve terminals that release more neurotransmitter per impulse have a higher incidence of synapses with more than one active zone, and this is correlated with more calcium build-up during stimulation. However, comparison of synaptic structure in neurons with different physiological phenotypes indicates no major differences in structure that could account for their different levels of neurotransmitter release per impulse, and release per synapse differs among neurons despite similar calcium build-up in their terminals during stimulation. The evidence indicates differences in calcium sensitivity of the release process among neurons as an aspect of physiological specialization.

Functional expression of the rat pancreatic islet glucose-dependent insulinotropic polypeptide receptor: ligand binding and intracellular signaling properties.

Incretins are endogenous peptides released from the gastrointestinal tract into the circulation during a meal that potentiate glucose-stimulated insulin secretion. At present, there are two established incretins: glucose-dependent insulinotropic polypeptide (GIP) and the truncated glucagon-like peptides (tGLPs), which are now being investigated for use in the treatment of diabetes mellitus. In the present study we cloned a rat islet GIP receptor complementary DNA (GIP-R1) to answer several important questions regarding the ligand-binding and intracellular signaling properties of the GP receptor. GIP-R1, when expressed transiently in monkey kidney (COS-7) or stably in Chinese hamster ovary (CHO-K1) cells, demonstrated comparable high affinity binding for either synthetic porcine (sp) GIP or synthetic human (sh) GIP. The IC50 values for displacement of [125I]spGIP in CHO-K1 cells were 2.6 +/- 0.8 and 3.1 +/- 0.9 nM for two different preparations of shGIP, and 3.7 +/- 1.5 and 3.6 +/- 0.4 nM for two preparations of spGIP. Saturation isotherms obtained with both intact cells and membranes gave monophasic binding curves with apparent Kd values of 204 +/- 17 and 334 +/- 94 pM, respectively. Cells expressed 12-15 x 10(3) receptors/cell. In COS-7 cells, spGIP and shGIP also exhibited similar IC50 values (7.6 +/- 1.2 and 8.9 +/- 1.8 nM, respectively). The receptor in CHO-K1 cells bound GIP-(1-30) with lower affinity (IC50 = 39 +/- 17 nM), whereas the fragments GIP-(19-30), GIP-(18-28), and GIP-(21-26) showed no apparent binding. The specificity of the receptor was further examined using several structurally related peptides. Surprisingly, exendin-(9-39) [Ex-(9-39)], a GLP-1 receptor antagonist, and Ex-4-(1-39), a GLP-1 receptor agonist, demonstrated some affinity for the GIP receptor, with 39% and 21% displacement of [125I]spGIP, respectively, at 1 microM. Other members of the secretin/vasoactive intestinal peptide family of peptides tested showed no interaction. GIP-R1 receptor binding correlated with activation of the adenylyl cyclase system, whereby spGIP and shGIP evoked concentration-dependent increases in cAMP accumulation with EC50 values of 8.7 +/- 1.5 x 10(-10)M and 8.1 +/- 1.6 x 10(-10)M for spGIP and shGIP, respectively. Increases in cAMP in the presence of 10 nM spGIP were not dependent on the ambient glucose concentration, with 22- and 18-fold increases in cAMP accumulation at 0.1 and 5.5 mM glucose, respectively.(ABSTRACT TRUNCATED AT 400 WORDS)

Synaptic regulation of glial protein expression in vivo.

We investigated signaling between individual nerve terminals and perisynaptic Schwann cells, the teloglial cells that cover neuromuscular junctions. When deprived of neuronal activity in vivo, either by motor nerve transection or tetrodotoxin injection, perisynaptic Schwann cells rapidly up-regulated glial fibrillary acidic protein. Addition of transcription or translation inhibitors to excised muscles prevented this increase. Stimulation of cut nerves prevented glial fibrillary acidic protein increases even when postsynaptic nicotinic receptors were blocked, but not when neurotransmitter release was blocked with omega-conotoxin GVIA. We conclude that there is a nerve terminal to glial signal, requiring presynaptic neurotransmitter release, which regulates perisynaptic Schwann cell genes. This may be a general principle since many types of glial are sensitive to transmitters applied in vitro or released in situ.