Age-related effects of Ginkgo biloba extract on synaptic plasticity and excitability

EGb 761 is a standardized extract from the Ginkgo biloba leaf and is purported to improve age-related memory impairment. The acute and chronic effect of EGb 761 on synaptic transmission and plasticity in hippocampal slices from young adult (8-12 weeks) and aged (18-24 months) C57B1/6 mice was tested because hippocampal plasticity is believed to be a key component of memory. Acutely applied EGb 761 significantly increased neuronal excitability in slices from aged mice by reducing the population spike threshold and increased the early phase of long-term potentiation, though there was no effect in slices from young adults. In chronically treated mice fed for 30 days with an EGb 761-supplemented diet, EGb 761 significantly increased the population spike threshold and long-term potentiation in slices from aged animals, but had no effect on slices from young adults. The rapid effects of EGb 761 on plasticity indicate a direct interaction with the glutamatergic system and raise interesting implications with respect to a mechanism explaining its effect on cognitive enhancement in human subjects experiencing dementia. (C) 2003 Elsevier Inc. All rights reserved.

The interactions of flavonoids within neuronal signalling pathways

Emerging evidence suggests that dietary phytochemicals, in particular flavonoids, may exert beneficial effects in the central nervous system by protecting neurons against stress-induced injury, by suppressing neuroinflammation and by promoting neurocognitive performance, through changes in synaptic plasticity. It is likely that flavonoids exert such effects in neurons, through selective actions on different components within a number of protein kinase and lipid kinase signalling cascades, such as phosphatidylinositol-3 kinase (PI3K)/Akt, protein kinase C and mitogen-activated protein kinase. This review details the potential inhibitory or stimulatory actions of flavonoids within these pathways, and describes how such interactions are likely to affect cellular function through changes in the activation state of target molecules and/or by modulating gene expression. Although, precise sites of action are presently unknown, their abilities to: (1) bind to ATP binding sites on enzymes and receptors; (2) modulate the activity of kinases directly; (3) affect the function of important phosphatases; (4) preserve neuronal Ca2+ homeostasis; and (5) modulate signalling cascades lying downstream of kinases, are explored. Future research directions are outlined in relation to their precise site(s) of action within the signalling pathways and the sequence of events that allow them to regulate neuronal function in the central nervous system.

The Ca(v)2.3 calcium channel antagonist SNX-482 reduces dorsal horn neuronal responses in a rat model of chronic neuropathic pain

Neuropathic pain is a difficult state to treat, characterized by alterations in sensory processing that can include allodynia (touch-evoked pain). Evidence exists for nerve damage-induced plasticity in both transmission and modulatory systems, including changes in voltage-dependent calcium channel (VDCC) expression and function; however, the role of Ca(v)2.3 calcium channels has not clearly been defined. Here, the effects of SNX-482, a selective Ca(v)2.3 antagonist, on sensory transmission at the spinal cord level have been investigated in the rat. The spinal nerve ligation (SNL) model of chronic neuropathic pain [Kim & Chung, (1992) Pain, 50, 355-363] was used to induce mechanical allodynia, as tested on the ipsilateral hindpaw. In vivo electrophysiological measurements of dorsal horn neuronal responses to innocuous and noxious electrical and natural stimuli were made after SNL and compared to sham-operated animals. Spinal SNX-482 (0.5-4 mu g/50 mu L) exerted dose-related inhibitions of noxious C-fibre- and A delta-fibre-mediated neuronal responses in conditions of neuropathy, but not in sham-operated animals. Measures of spinal cord hyperexcitability and nociception were most susceptible to SNX-482. In contrast, non-noxious A beta-mediated responses were not affected by SNX-482. Moreover, responses to innocuous mechanical and also thermal stimuli were more sensitive to SNX-482 in SNL than control animals. This study is the first to demonstrate an antinociceptive role for SNX-482-sensitive channels in dorsal horn neurons during neuropathy. These data are consistent with plasticity in Ca(V)2.3 calcium channel expression and suggest a potential selective target to reduce nociceptive transmission during conditions of nerve damage.

Endogenous cholinergic tone modulates spontaneous network level neuronal activity in primary cortical cultures grown on multi-electrode arrays

Background Cortical cultures grown long-term on multi-electrode arrays (MEAs) are frequently and extensively used as models of cortical networks in studies of neuronal firing activity, neuropharmacology, toxicology and mechanisms underlying synaptic plasticity. However, in contrast to the predominantly asynchronous neuronal firing activity exhibited by intact cortex, electrophysiological activity of mature cortical cultures is dominated by spontaneous epileptiform-like global burst events which hinders their effective use in network-level studies, particularly for neurally-controlled animat (‘artificial animal’) applications. Thus, the identification of culture features that can be exploited to produce neuronal activity more representative of that seen in vivo could increase the utility and relevance of studies that employ these preparations. Acetylcholine has a recognised neuromodulatory role affecting excitability, rhythmicity, plasticity and information flow in vivo although its endogenous production by cortical cultures and subsequent functional influence upon neuronal excitability remains unknown. Results Consequently, using MEA electrophysiological recording supported by immunohistochemical and RT-qPCR methods, we demonstrate for the first time, the presence of intrinsic cholinergic neurons and significant, endogenous cholinergic tone in cortical cultures with a characterisation of the muscarinic and nicotinic components that underlie modulation of spontaneous neuronal activity. We found that tonic muscarinic ACh receptor (mAChR) activation affects global excitability and burst event regularity in a culture age-dependent manner whilst, in contrast, tonic nicotinic ACh receptor (nAChR) activation can modulate burst duration and the proportion of spikes occurring within bursts in a spatio-temporal fashion. Conclusions We suggest that the presence of significant endogenous cholinergic tone in cortical cultures and the comparability of its modulatory effects to those seen in intact brain tissues support emerging, exploitable commonalities between in vivo and in vitro preparations. We conclude that experimental manipulation of endogenous cholinergic tone could offer a novel opportunity to improve the use of cortical cultures for studies of network-level mechanisms in a manner that remains largely consistent with its functional role.

Formation of cortical plasticity in older adults following tDCS and motor training

Neurodegeneration accompanies the process of natural aging, reducing the ability to perform functional daily activities. Transcranial direct current stimulation (tDCS) alters neuronal excitability and motor performance; however its beneficial effect on the induction of primary motor cortex (M1) plasticity in older adults is unclear. Moreover, little is known as to whether the tDCS electrode arrangement differentially affects M1 plasticity and motor performance in this population. In a double-blinded, cross-over trial, we compared unilateral, bilateral and sham tDCS combined with visuomotor tracking, on M1 plasticity and motor performance of the non-dominant upper limb, immediately post and 30 min following stimulation. We found (a) unilateral and bilateral tDCS decreased tracking error by 12–22% at both time points; with sham decreasing tracking error by 10% at 30 min only, (b) at both time points, motor evoked potentials (MEPs) were facilitated (38–54%) and short-interval intracortical inhibition was released (21–36%) for unilateral and bilateral conditions relative to sham, (c) there were no differences between unilateral and bilateral conditions for any measure. These findings suggest that tDCS modulated elements of M1 plasticity, which improved motor performance irrespective of the electrode arrangement. The results provide preliminary evidence indicating that tDCS is a safe non-invasive tool to preserve or improve neurological function and motor control in older adults.

Targeting neuronal populations by AAV-mediated gene transfer for studying the endocannabinoid system

The cannabinoid type 1 (CB1) receptor is involved in a plethora of physiological functions and heterogeneously expressed on different neuronal populations. Several conditional loss-of-function studies revealed distinct effects of CB1 receptor signaling on glutamatergic and GABAergic neurons, respectively. To gain a comprehensive picture of CB1 receptor-mediated effects, the present study aimed at developing a gain-of-function approach, which complements conditional loss-of-function studies. Therefore, adeno-associated virus (AAV)-mediated gene delivery and Cre-mediated recombination were combined to recreate an innovative method, which ensures region- and cell type-specific transgene expression in the brain. This method was used to overexpress the CB1 receptor in glutamatergic pyramidal neurons of the mouse hippocampus. Enhanced CB1 receptor activity at glutamatergic terminals caused impairment in hippocampus-dependent memory performance. On the other hand, elevated CB1 receptor levels provoked an increased protection against kainic acid-induced seizures and against excitotoxic neuronal cell death. This finding indicates the protective role of CB1 receptor on hippocampal glutamatergic terminals as a molecular stout guard in controlling excessive neuronal network activity. Hence, CB1 receptor on glutamatergic hippocampal neurons may represent a target for novel agents to restrain excitotoxic events and to treat neurodegenerative diseases. Endocannabinoid synthesizing and degrading enzymes tightly regulate endocannabinoid signaling, and thus, represent a promising therapeutic target. To further elucidate the precise function of the 2-AG degrading enzyme monoacylglycerol lipase (MAGL), MAGL was overexpressed specifically in hippocampal pyramidal neurons. This genetic modification resulted in highly increased MAGL activity accompanied by a 50 % decrease in 2-AG levels without affecting the content of arachidonic acid and anandamide. Elevated MAGL protein levels at glutamatergic terminals eliminated depolarization-induced suppression of excitation (DSE), while depolarization-induced suppression of inhibition (DSI) was unchanged. This result indicates that the on-demand availability of the endocannabinoid 2-AG is crucial for short-term plasticity at glutamatergic synapses in the hippocampus. Mice overexpressing MAGL exhibited elevated corticosterone levels under basal conditions and an increase in anxiety-like behavior, but surprisingly, showed no changes in aversive memory formation and in seizure susceptibility. This finding suggests that 2 AG-mediated hippocampal DSE is essential for adapting to aversive situations, but is not required to form aversive memory and to protect against kainic acid-induced seizures. Thus, specific inhibition of MAGL expressed in hippocampal pyramidal neurons may represent a potential treatment strategy for anxiety and stress disorders. Finally, the method of AAV-mediated cell type-specific transgene expression was advanced to allow drug-inducible and reversible transgene expression. Therefore, elements of the tetracycline-controlled gene expression system were incorporated in our “conditional” AAV vector. This approach showed that transgene expression is switched on after drug application and that background activity in the uninduced state was only detectable in scattered cells of the hippocampus. Thus, this AAV vector will proof useful for future research applications and gene therapy approaches.

Spatio-Temporal Credit Assignment in Neuronal Population Learning

n learning from trial and error, animals need to relate behavioral decisions to environmental reinforcement even though it may be difficult to assign credit to a particular decision when outcomes are uncertain or subject to delays. When considering the biophysical basis of learning, the credit-assignment problem is compounded because the behavioral decisions themselves result from the spatio-temporal aggregation of many synaptic releases. We present a model of plasticity induction for reinforcement learning in a population of leaky integrate and fire neurons which is based on a cascade of synaptic memory traces. Each synaptic cascade correlates presynaptic input first with postsynaptic events, next with the behavioral decisions and finally with external reinforcement. For operant conditioning, learning succeeds even when reinforcement is delivered with a delay so large that temporal contiguity between decision and pertinent reward is lost due to intervening decisions which are themselves subject to delayed reinforcement. This shows that the model provides a viable mechanism for temporal credit assignment. Further, learning speeds up with increasing population size, so the plasticity cascade simultaneously addresses the spatial problem of assigning credit to synapses in different population neurons. Simulations on other tasks, such as sequential decision making, serve to contrast the performance of the proposed scheme to that of temporal difference-based learning. We argue that, due to their comparative robustness, synaptic plasticity cascades are attractive basic models of reinforcement learning in the brain.

Extremes of Lineage Plasticity in the Drosophila Brain

An often-overlooked aspect of neural plasticity is the plasticity of neuronal composition, in which the numbers of neurons of particular classes are altered in response to environment and experience. The Drosophila brain features several well-characterized lineages in which a single neuroblast gives rise to multiple neuronal classes in a stereotyped sequence during development. We find that in the intrinsic mushroom body neuron lineage, the numbers for each class are highly plastic, depending on the timing of temporal fate transitions and the rate of neuroblast proliferation. For example, mushroom body neuroblast cycling can continue under starvation conditions, uncoupled from temporal fate transitions that depend on extrinsic cues reflecting organismal growth and development. In contrast, the proliferation rates of antennal lobe lineages are closely associated with organismal development, and their temporal fate changes appear to be cell-cycle dependent, such that the same numbers and types of uniglomerular projection neurons innervate the antennal lobe following various perturbations. We propose that this surprising difference in plasticity for these brain lineages is adaptive, given their respective roles as parallel processors versus discrete carriers of olfactory information.

Extremes of Lineage Plasticity in the Drosophila Brain

An often-overlooked aspect of neural plasticity is the plasticity of neuronal composition, in which the numbers of neurons of particular classes are altered in response to environment and experience. The Drosophila brain features several well-characterized lineages in which a single neuroblast gives rise to multiple neuronal classes in a stereotyped sequence during development [1]. We find that in the intrinsic mushroom body neuron lineage, the numbers for each class are highly plastic, depending on the timing of temporal fate transitions and the rate of neuroblast proliferation. For example, mushroom body neuroblast cycling can continue under starvation conditions, uncoupled from temporal fate transitions that depend on extrinsic cues reflecting organismal growth and development. In contrast, the proliferation rates of antennal lobe lineages are closely associated with organismal development, and their temporal fate changes appear to be cell cycle-dependent, such that the same numbers and types of uniglomerular projection neurons innervate the antennal lobe following various perturbations. We propose that this surprising difference in plasticity for these brain lineages is adaptive, given their respective roles as parallel processors versus discrete carriers of olfactory information.

Functional, structural and molecular plasticity of mammalian skeletal muscle in response to exercise stimuli

Biological systems have acquired effective adaptive strategies to cope with physiological challenges and to maximize biochemical processes under imposed constraints. Striated muscle tissue demonstrates a remarkable malleability and can adjust its metabolic and contractile makeup in response to alterations in functional demands. Activity-dependent muscle plasticity therefore represents a unique model to investigate the regulatory machinery underlying phenotypic adaptations in a fully differentiated tissue. Adjustments in form and function of mammalian muscle have so far been characterized at a descriptive level, and several major themes have evolved. These imply that mechanical, metabolic and neuronal perturbations in recruited muscle groups relay to the specific processes being activated by the complex physiological stimulus of exercise. The important relationship between the phenotypic stimuli and consequent muscular modifications is reflected by coordinated differences at the transcript level that match structural and functional adjustments in the new training steady state. Permanent alterations of gene expression thus represent a major strategy for the integration of phenotypic stimuli into remodeling of muscle makeup. A unifying theory on the molecular mechanism that connects the single exercise stimulus to the multi-faceted adjustments made after the repeated impact of the muscular stress remains elusive. Recently, master switches have been recognized that sense and transduce the individual physical and chemical perturbations induced by physiological challenges via signaling cascades to downstream gene expression events. Molecular observations on signaling systems also extend the long-known evidence for desensitization of the muscle response to endurance exercise after the repeated impact of the stimulus that occurs with training. Integrative approaches involving the manipulation of single factors and the systematic monitoring of downstream effects at multiple levels would appear to be the ultimate method for pinpointing the mechanism of muscle remodeling. The identification of the basic relationships underlying the malleability of muscle tissue is likely to be of relevance for our understanding of compensatory processes in other tissues, species and organisms.

Plasticity in the adult language system: a longitudinal electrophysiological study on second language learning

Event-related potentials (ERPs) were used to trace changes in brain activity related to progress in second language learning. Twelve English-speaking exchange students learning German in Switzerland were recruited. ERPs to visually presented single words from the subjects' native language (English), second language (German) and an unknown language (Romansh) were measured before (day 1) and after (day 2) 5 months of intense German language learning. When comparing ERPs to German words from day 1 and day 2, we found topographic differences between 396 and 540 ms. These differences could be interpreted as a latency shift indicating faster processing of German words on day 2. Source analysis indicated that the topographic differences were accounted for by shorter activation of left inferior frontal gyrus (IFG) on day 2. In ERPs to English words, we found Global Field Power differences between 472 and 644 ms. This may due to memory traces related to English words being less easily activated on day 2. Alternatively, it might reflect the fact that--with German words becoming familiar on day 2--English words loose their oddball character and thus produce a weaker P300-like effect on day 2. In ERPs to Romansh words, no differences were observed. Our results reflect plasticity in the neuronal networks underlying second language acquisition. They indicate that with a higher level of second language proficiency, second language word processing is faster and requires shorter frontal activation. Thus, our results suggest that the reduced IFG activation found in previous fMRI studies might not reflect a generally lower activation but rather a shorter duration of activity.

Lobe specific Ca2+-calmodulin nano-domain in neuronal spines: a single molecule level analysis.

Calmodulin (CaM) is a ubiquitous Ca(2+) buffer and second messenger that affects cellular function as diverse as cardiac excitability, synaptic plasticity, and gene transcription. In CA1 pyramidal neurons, CaM regulates two opposing Ca(2+)-dependent processes that underlie memory formation: long-term potentiation (LTP) and long-term depression (LTD). Induction of LTP and LTD require activation of Ca(2+)-CaM-dependent enzymes: Ca(2+)/CaM-dependent kinase II (CaMKII) and calcineurin, respectively. Yet, it remains unclear as to how Ca(2+) and CaM produce these two opposing effects, LTP and LTD. CaM binds 4 Ca(2+) ions: two in its N-terminal lobe and two in its C-terminal lobe. Experimental studies have shown that the N- and C-terminal lobes of CaM have different binding kinetics toward Ca(2+) and its downstream targets. This may suggest that each lobe of CaM differentially responds to Ca(2+) signal patterns. Here, we use a novel event-driven particle-based Monte Carlo simulation and statistical point pattern analysis to explore the spatial and temporal dynamics of lobe-specific Ca(2+)-CaM interaction at the single molecule level. We show that the N-lobe of CaM, but not the C-lobe, exhibits a nano-scale domain of activation that is highly sensitive to the location of Ca(2+) channels, and to the microscopic injection rate of Ca(2+) ions. We also demonstrate that Ca(2+) saturation takes place via two different pathways depending on the Ca(2+) injection rate, one dominated by the N-terminal lobe, and the other one by the C-terminal lobe. Taken together, these results suggest that the two lobes of CaM function as distinct Ca(2+) sensors that can differentially transduce Ca(2+) influx to downstream targets. We discuss a possible role of the N-terminal lobe-specific Ca(2+)-CaM nano-domain in CaMKII activation required for the induction of synaptic plasticity.

Role of entorhinal cortical plasticity in spatial and contextual memory

It is well accepted that the hippocampus (HIP) is important for spatial and contextual memories, however, it is not clear if the entorhinal cortex (EC), the main input/output structure for the hippocampus, is also necessary for memory storage. Damage to the EC in humans results in memory deficits. However, animal studies report conflicting results on whether the EC is necessary for spatial and contextual memory. Memory consolidation requires gene expression and protein synthesis, mediated by signaling cascades and transcription factors. Extracellular-signal regulated kinase (ERK) cascade activity is necessary for long-term memory in several tasks, including those that test spatial and contextual memory. In this work, we explore the role of ERK-mediated plasticity in the EC on spatial and contextual memory. ^ To evaluate this role, post-training infusions of reversible pharmacological inhibitors specific for the ERK cascade that do not affect normal neuronal activity were targeted directly to the EC of awake, behaving animals. This technique provides spatial and temporal control over the inhibition of the ERK cascade without affecting performance during training or testing. Using the Morris water maze to study spatial memory, we found that ERK inhibition in the EC resulted in long-term memory deficits consistent with a loss of spatial strategy information. When animals were allowed to learn and consolidate a spatial strategy for solving the task prior to training and ERK inhibition, the deficit was alleviated. To study contextual memory, we trained animals in a cued fear-conditioning task and saw an increase in the activation of ERK in the EC 90 minutes following training. ERK inhibition in the EC over this time point, but not at an earlier time point, resulted in increased freezing to the context, but not to the tone, during a 48-hour retention test. In addition, animals froze maximally at the time the shock was given during training; similar to naïve animals receiving additional training, suggesting that ERK-mediated plasticity in the EC normally suppresses the temporal nature of the freezing response. These findings demonstrate that plasticity in the EC is necessary for both spatial and contextual memory, specifically in the retention of behavioral strategies. ^

TURNOVER RATE OF THE NEURONAL CONNEXIN CX36 IN HELA CELLS

Electrical synapses formed of the gap junction protein Cx36 show a great deal of functional plasticity, much dependent on changes in phosphorylation state of the connexin. However, gap junction turnover may also be important for regulating cell-cell communication, and turnover rates of Cx36 have not been studied. Connexins have relatively fast turnover rates, with short half-lives measured to be 1.5 to 3.5 hours in pulse-chase analyses of connexins (Cx26 and Cx43) in tissue culture cells and whole organs. We utilized HaloTag technology to study the turnover rate of Cx36 in transiently transfected HeLa cells. The HaloTag protein forms irreversible covalent bonds with chloroalkane ligands, allowing pulse-chase experiments to be performed very specifically. The HaloTag open reading frame was inserted into an internal site in the C-terminus of Cx36 designed not to disrupt the regulatory phosphorylation sites and not to block the C-terminal PDZ interaction motif. Functional properties of Cx36-Halo were assessed by Neurobiotin tracer coupling, live cell imaging, and immunostaining. For the pulse-chase study, transiently transfected HeLa cells were pulse labeled with Oregon Green (OG) HaloTag ligand and chase labeled at various times with tetramethylrhodamine (TMR) HaloTag ligand. Cx36-Halo formed large junctional plaques at sites of contact between transfected HeLa cells and was also contained in a large number of intracellular vesicles. The Cx36-Halo transfected HeLa cells supported Neurobiotin tracer coupling that was regulated by activation and inhibition of PKA in the same manner as wild-type Cx36 transfected cells. In the pulse-chase study, junctional protein labeled with the pulse ligand (OG) was gradually replaced by newly synthesized Cx36 labeled with the chase ligand (TMR). The half-life for turnover of protein in junctional plaques was 2.8 hours. Treatment of the pulse-labeled cells with Brefeldin A (BFA) prevented the addition of new connexins to junctional plaques, suggesting that the assembly of Cx36 into gap junctions involves the traditional ER-Golgi-TGN-plasma membrane pathway. In conclusion, Cx36-Halo is functional and has a turnover rate in HeLa cells similar to that of other connexins that have been studied. This turnover rate is likely too slow to contribute substantially to short-term changes in coupling of neurons driven by transmitters such as dopamine, which take minutes to achieve. However, turnover may contribute to longer-term changes in coupling.

Context-sensitive synaptic plasticity and temporal-to-spatial transformations in hippocampal slices

Hippocampal slices are used to show that, as a temporal input pattern of activity flows through a neuronal layer, a temporal-to-spatial transformation takes place. That is, neurons can respond selectively to the first or second of a pair of input pulses, thus transforming different temporal patterns of activity into the activity of different neurons. This is demonstrated using associative long-term potentiation of polysynaptic CA1 responses as an activity-dependent marker: by depolarizing a postsynaptic CA1 neuron exclusively with the first or second of a pair of pulses from the dentate gyrus, it is possible to “tag” different subpopulations of CA3 neurons. This technique allows sampling of a population of neurons without recording simultaneously from multiple neurons. Furthermore, it reflects a biologically plausible mechanism by which single neurons may develop selective responses to time-varying stimuli and permits the induction of context-sensitive synaptic plasticity. These experimental results support the view that networks of neurons are intrinsically able to process temporal information and that it is not necessary to invoke the existence of internal clocks or delay lines for temporal processing on the time scale of tens to hundreds of milliseconds.