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Die Vorgaben von Leitlinien erweisen sich haufig als nicht praxistauglich oder spiegeln nur ein verzerrtes Bild des Praxisalltags wider. „Klug entscheiden in der Schmerzmedizin“ bedeutet aber nicht nur, externe, sondern auch interne Evidenz sowie die Patientenpraferenz zu berucksichtigen. Eine Arbeitsgruppe der DGS hat sich intensiv mit der Frage beschaftigt, wie sich Leitlinien in der Versorgung von Patienten mit chronischen Schmerzen auswirken und auswirken durfen.

Background The term 'visually induced analgesia' describes a reduced pain perception induced by watching the painful body part as opposed to watching a neutral object. In chronic back pain patients, experimental pain, movement-induced pain and habitual pain can be reduced with visual feedback. Visual feedback can also enhance the effects of both massage treatment and manual therapy. The impact of somatosensory attentional processes remains unclear. Methods In the current study, participants received painful electrical stimuli to their thumb and back while being presented with either a real-time video of their thumb or back (factor feedback). In addition, using an oddball paradigm, they had to count the number of deviant stimuli, applied to either their back or thumb (factor attention) and rate the pain intensity. Results We found a significant main effect for attention with decreased pain ratings during attention. There was no main effect for visual feedback and no significant interaction between visual feedback and attention. Post-hoc tests revealed that the lowest pain intensity ratings were achieved during visual feedback of the back/ thumb and counting at the back/ thumb. Conclusion These data suggest that the modulation of perceived acute pain by visually induced analgesia may be influenced by a simultaneous somatosensory attention task. Significance Somatosensory attention reduced experimental pain intensity in the thumb and back in the presence of both congruent and incongruent visual feedback. We found no significant visual feedback effect on the complex interplay between visual feedback and somatosensory attention.

Opioid receptor agonists are known to relieve restless legs syndrome (RLS) symptoms, including both sensory and motor events, as well as improving sleep. The mechanisms of action of opioids in RLS are still a matter of speculation. The mechanisms by which endogenous opioids contribute to the pathophysiology of this polygenetic disorder, in which there are a number of variants, including developmental factors, remains unknown. A summary of the cellular mode of action of morphine and its (partial) antagonist naloxone via α-amino-3-hydroxy-5-methyl-4-isoxazolepropionic acid (AMPA) receptors and the involvement of dendritic spine activation is described. By targeting pain and its consequences, opioids are the first-line treatment in many diseases and conditions with both acute and chronic pain and have thus been used in both acute and chronic pain conditions over the last 40 years. Addiction, dependence, and tolerability of opioids show a wide variability interindividually, as the response to opioids is influenced by a complex combination of genetic, molecular, and phenotypic factors. Although several trials have now addressed opioid treatment in RLS, hyperalgesia as a complication of long-term opioid treatment, or opioid–opioid interaction have not received much attention so far. Therapeutic opioids may act not only on opioid receptors but also via histamine or N -methyl-d-aspartate (NMDA) receptors. In patients with RLS, one of the few studies investigating opioid bindings found that possible brain regions involved in the severity of RLS symptoms are similar to those known to be involved in chronic pain, such as the medial pain system (medial thalamus, amygdala, caudate nucleus, anterior cingulate gyrus, insular cortex, and orbitofrontal cortex). The results of this diprenorphine positron emission tomography study suggested that the more severe the RLS, the greater the release of endogenous opioids. Since 1993, when the first small controlled study was performed with oxycodone in RLS, opioids have been considered an efficacious off-label therapy in patients with severe RLS. A recent trial has proved the efficacy of a combination of prolonged release oxycodone/naloxone in patients with severe RLS as second-line therapy, with a mean dosage of 10/5 mg twice daily (mean difference of International Restless Legs Syndrome Study Group Rating Scale (IRLS) score between groups at 12 weeks: 8.15), and has now been licensed as the first opioid therapy in Europe. The current results from both short- and long-term trials and studies with opioids encourage optimism in alleviating RLS symptoms in patients with severe RLS, or possibly during or after augmentation.

Background Visual analgesia refers to the phenomena where people report decreased pain intensity when they see the painful or painfully stimulated body part. Alongside pain, sensorimotor impairment (i.e., disturbed proprioception) is also evident in chronic pain. This study aims to investigate whether real-time visual feedback offers additional pain relief and proprioceptive improvement when used in combination with recommended therapies in neck pain patients who received manual therapy with or without real-time visual feedback. Methods A total of 29 neck pain patients were recruited in an outpatient physical therapy practice. Patients were randomly allocated to receive manual therapy of the cervical spine with real-time visual feedback or to a control group where patients received manual therapy without real-time visual feedback. Habitual pain intensity, the pressure pain threshold at the zygapophyseal joint of C2-C3 and the superior angle of the scapulae and cervical proprioception were assessed before and immediately after the intervention by a blinded assessor. Results A between-group comparison revealed a significant reduction in habitual pain in the real-time visual feedback group. No differences were found for the pressure pain threshold or proprioceptive performance. Conclusions Real-time visual feedback combined with manual therapy enhanced the analgesic effect of manual therapy in neck pain patients, but had no positive effect on the pressure pain threshold and cervical joint position sense. The technical demands for integrating real-time visual feedback into daily practice to reduce habitual pain are low, have low costs and are easy to apply. Significance Real-time visual feedback reduces habitual pain immediately after the intervention. Due to its easy integration, it may be an effective adjunct to recommended interventions (i.e., manual therapy) in patients with neck pain.

Background Tramadol and tilidine (in combination with naloxone) are used as weak opioid analgesics in Germany. Tramadol is not scheduled in the German Narcotic Drugs Act. Tilidine is scheduled, whereas Tilidine in fixed combinations with naloxone is exempt from some of the provisions of the Narcotic Drugs Act. Recent reports on misuse of both substances led to an evaluation of their potential for misuse, abuse, and dependency by the expert advisory committee established by the German Federal Government, resident at the Federal Institute for Drugs and Medical Devices. Methods A subcommittee formulated key questions and identified available data sources for each of these questions. Additional information was solicited where necessary, including a survey among a panel of pharmacists, a survey in an addiction clinic, analysis of prescription patterns, and information from the boards of pharmacists of the federal states and the Federal Bureau of Criminal Investigation. Results Analgesic efficiency in the treatment of acute and chronic pain has been proven for both tramadol and tilidine/naloxone. For tramadol, high evidence has been confirmed in systematic reviews, and tramadol is listed in national and international guidelines on acute and chronic pain management. Animal and human studies found a low potential for misuse, abuse, and dependency for both substances. Information from 2 tramadol safety databases allowed calculation of the incidence of abuse or dependency as 0.21 and 0.12 cases per million defined daily dosages (DDDs), with lower incidences in recent years. For tilidine/naloxone, the incidence was calculated as 0.43 cases per million DDDs for oral solution and 0.18 for slow-release tablets. In an online survey among German pharmacies as well as in the reports from state pharmacy boards, fraud attempts were repeated more frequently with tilidine/naloxone than with tramadol in the last 2 years. The Federal Bureau of Criminal Investigations reported prescription fraud only with tilidine/naloxone and predominantly in the region of Berlin. Dependency on tramadol or tilidine/naloxone is reported only rarely from addiction counseling centers. One third of the patients surveyed in an addiction clinic reported experiences with tramadol or tilidine/naloxone, but mostly with duration of less than 4 weeks and with a medical prescription based on a reasonable indication. Also, occasional illegal use of opioid analgesics as a substitute of heroin was reported. An evaluation of pooled data from statutory health insurance companies found 2.5% of persons receiving at least 1 prescription of tramadol or the combination of tilidine and naloxone in 2009 (1.6% with tramadol and 1.0% with tilidine/naloxone). High usage with more than 180 DDDs per year was found in 8.6% of patients treated with tramadol and 17.2% of patients with tilidine/naloxone. Conclusions In conclusion, the subcommittee of the expert advisory committee found a low potential for misuse, abuse, and dependency for tramadol, and a low prevalence in clinical practice. Considerable less information is available for the combination of tilidine and naloxone. However, the cumulation of evidence indicated a higher risk of misuse, abuse, and dependency for tilidine/naloxone solution, but not for slow-release tablets.

Brain-derived steroids (neurosteroids) constitute a new group of steroids synthesized in brain cells independently from peripheral sources. While the membraneous actions of these steroids have been characterized in some detail, their genomic effects are less well studied up to now. In the present study, the action of the neurosteroids dehydroepiandrosterone, pregnenolone and pregnanolone on proopiomelanocortin (POMC)-gene expression was investigated in pituitary AtT 20 cells, which were cultured under serum-free conditions and at strictly controlled steroid levels. In these cells, POMC-gene expression is known to be negatively regulated by adrenocortical glucocorticoids. Quantitative Northern blot analysis showed that none of these neurosteroids or their conjugates affected POMC-mRNA levels even at the highest concentrations employed. The 21-OH steroid corticosterone was used as a control and markedly suppressed the synthesis of POMC-mRNA. These results suggest that neurosteroids do not control POMC-gene expression at the level of the pituitary corticotroph and exclude significant effects of these compounds at the glucocorticoid receptor.

Postoperative cognitive dysfunction (POCD) is a decline in cognitive performance after a surgery performed under anaesthesia. The exact roles of surgery and/or anaesthesia for facilitating POCD are unclear. This study investigates the effects of isoflurane anaesthesia on cognitive performance and cellular mechanisms involved in learning and memory function. Male C57BL6/J mice (age: 4-5 months) were anaesthetized with isoflurane in oxygen/air (FiO(2)=0.5) for 2h, non-anaesthetized mice served as controls. After 24h, neurocognitive function, in vitro long-term potentiation (LTP), or protein expression were evaluated. In a visuospatial test, anaesthetized mice showed better cognitive performance as they learned faster compared to controls. In hippocampal slices of anaesthetized mice, in vitro LTP was enhanced as reflected in an increased extracellular field potential (fEPSP) slope after 1h to 210.2+/-17% (control: 156.8+/-7.2%; n=14; p

Background The neuronal and molecular targets of the inhalational general anesthetic xenon are a matter of debate. The current knowledge is largely based on studies using neurons in culture or heterologous expression systems. In the current study, the authors evaluated for the first time the effect of xenon on synaptic transmission in the basolateral amygdala in an in vitro brain slice preparation of the mouse. Methods A patch clamp technique was used to evaluate the effects of xenon on N-methyl-d-aspartate (NMDA) and alpha-amino-3-hydroxy-5-methyl-4-isoxazolepropionic acid (AMPA) receptor-mediated excitatory postsynaptic currents (EPSCs), as well as on gamma-aminobutyric acid type A receptor-mediated inhibitory postsynaptic currents. The currents were either evoked upon electrical stimulation (NMDA-eEPSCs, AMPA-eEPSCs) or upon focal, laser-guided photolysis of caged l-glutamate (p-NMDA-Cs, p-AMPA-Cs). In addition, the authors investigated the effects of xenon on miniature EPSCs. Results Xenon reversibly reduced basal synaptic transmission but had no effect on gamma-aminobutyric acid type A receptor-mediated inhibitory synaptic transmission. Xenon concentration-dependently diminished NMDA-eEPSCs and p-NMDA-Cs to the same amount. Likewise, xenon-induced reduction of AMPA-eEPSCs and p-AMPA-Cs did not differ. Xenon did not affect the frequency of miniature EPSCs but reduced their amplitude. Conclusions In the current study, xenon considerably depressed NMDA and AMPA receptor-mediated synaptic transmission in the basolateral amygdala without affecting inhibitory synaptic transmission. The results provide evidence that the effects of xenon on NMDA- and AMPA-EPSCs are primarily mediated via postsynaptic mechanisms.

Excessive N-methyl-D-aspartate (NMDA) receptor activation is widely accepted to mediate calcium-dependent glutamate excitotoxicity. The uncompetitive, voltage-dependent NMDA receptor antagonist memantine has been successfully used clinically in the treatment of neurodegenerative dementia and is internationally registered for the treatment of moderate to severe Alzheimer's disease. Glutamate release inhibitors (GRIs) may also be promising for the therapy of some neurodegenerative diseases. During the clinical use of GRIs, it could be questioned whether there would still be a sufficient number of active NMDA receptors to allow any additional effects of memantine or similar NMDA receptor antagonists. To address this question, we determined the fraction of NMDA receptors contributing to postsynaptic events in the presence of therapeutically relevant concentrations of the GRI riluzole (1 microM) using an in vitro hippocampal slice preparation. We measured the charge transfer of pharmacologically isolated excitatory synaptic responses before and after the application of the selective, competitive NMDA receptor antagonist D-AP5 (100 microM). The fraction of activated NMDA receptors under control conditions did not differ from those in the presence of riluzole. It is therefore likely that NMDA receptor antagonists would be able to exert additional therapeutic effects in combination therapy with GRIs.

Previously, we found that in the lateral amygdala (LA) of the mouse, WIN55,212-2 decreases both glutamatergic and GABAergic synaptic transmission via activation of the cannabinoid receptor type 1 (CB1), yet produces an overall reduction of neuronal excitability. This suggests that the effects on excitatory transmission override those on inhibitory transmission. Here we show that CB1 activation by WIN55,212-2 and Δ9-THC inhibits long-term depression (LTD) of basal synaptic transmission in the LA, induced by low-frequency stimulation (LFS; 900 pulses/1 Hz). The CB1 agonist WIN55,212-2 blocked LTD via Gi/o proteins, activation of inwardly rectifying K+ channels (Kirs), inhibition of the adenylate cyclase-protein kinase A (PKA) pathway, and PKA-dependent inhibition of voltage-gated N-type Ca2+ channels (N-type VGCCs). Interestingly, WIN55,212-2 effects on LTD were abolished in CB1 knock-out mice (CB1-KO), and in conditional mutants lacking CB1 expression only in GABAergic interneurons, but were still present in mutants lacking CB1 in principal forebrain neurons. LTD induction per se was unaffected by the CB1 antagonist SR141716A and was normally expressed in CB1-KO as well as in both conditional CB1 mutants. Our data demonstrate that activation of CB1 specifically located on GABAergic interneurons inhibits LTD in the LA. These findings suggest that CB1 expressed on either glutamatergic or GABAergic neurons play a differential role in the control of synaptic transmission and plasticity.

Visualizing entire neuronal networks for analysis in the intact brain has been impossible up to now. Techniques like computer tomography or magnetic resonance imaging (MRI) do not yield cellular resolution, and mechanical slicing procedures are insufficient to achieve high-resolution reconstructions in three dimensions. Here we present an approach that allows imaging of whole fixed mouse brains. We modified 'ultramicroscopy' by combining it with a special procedure to clear tissue. We show that this new technique allows optical sectioning of fixed mouse brains with cellular resolution and can be used to detect single GFP-labeled neurons in excised mouse hippocampi. We obtained three-dimensional (3D) images of dendritic trees and spines of populations of CA1 neurons in isolated hippocampi. Also in fruit flies and in mouse embryos, we were able to visualize details of the anatomy by imaging autofluorescence. Our method is ideally suited for high-throughput phenotype screening of transgenic mice and thus will benefit the investigation of disease models.

The gaseous anaesthetic N(2)O displays analgesic, anxiolytic, and amnesic properties and has addictive psychedelic effects. N(2)O can further act as a neuroprotective agent, but may also become neurotoxic under certain conditions. Here, we employed whole-cell patch-clamp techniques in acute brain slices, and electrical afferent and infrared-guided laser stimulation to examine how N(2)O (65%) can affect NMDA receptor (NMDAR)-mediated synaptic transmission to principal neurons (PNs) of the adult murine basolateral amygdala (BLA). The BLA plays a critical role in anaesthetic-induced amnesia, the formation of aversive memories, as well as in fear and addictive behaviour. We evoked NMDAR-mediated excitatory postsynaptic currents (NMDAR-EPSCs) in PNs of the BLA (BLA-PNs). We found these currents to be markedly decreased by N(2)O via pre- and postsynaptic actions: Without changing their kinetics and open probability, N(2)O impeded the voltage-dependent channel opening of NMDARs in BLA-PNs and diminished their unitary conductance as estimated by non-stationary fluctuation analysis. In addition, our data speak in favour of a N(2)O-produced reduction in the probability of glutamate release at the synapses generating the NMDAR-EPSCs. It is conceivable that these effects not only contribute to anaesthesia and anxiolysis, but also have bearings on learning and memory as well as excitotoxicity in the amygdala.

Background Chronic back pain (CBP) is a frequent debilitating and often treatment-resistant disorder. The awareness of one's own body seems to be essential in pain reduction through visual input. Visual feedback of the back reduces experimental pain perception in CBP at this site and watching the back during repeated lumbar spine movements reduces movement-evoked pain. In this study, we tested whether visual feedback alone can reduce habitual pain in CBP. Methods In a within-subject design, 19 CBP patients participated in an online visual feedback condition, watching one's own back. This was compared to several control conditions, such as watching a neutral object (book), a video of another person of the same sex, a picture of the own back, and keeping one's eyes closed in randomized order on five separate days. In each experimental session, participants rated habitual pain intensity and unpleasantness before and after the experimental manipulation. Results We present evidence that visual feedback by watching the site of chronic pain on a video screen alone is sufficient to reduce habitual chronic pain. No additional manipulation or movement was necessary. Conclusions These results suggest that online video feedback may be helpful in alleviating chronic pain.

AIM The aim of this study was to evaluate the role of the endogenous cannabinoid system in controlling neuroplasticity. METHODS The pain threshold for electrical stimuli was determined in transgenic mice lacking the cannabinoid receptor type 1 (CB1(-/-)) and in the corresponding respective wild-type animals. Electrophysiological experiments were performed in prepared brain slices to test the effect of endogenous and exogenous cannabinoids on synaptic transmission and long-term potentiation (LTP) in the amygdala. RESULTS The pain threshold was nearly identical in both groups for the first pain induction; however, with repeated pain induction it decreased to a significantly greater extent in the CB1(-/-) mice than in the wild-type animals. Synoptic transmission and the inducibility of LTP were not influenced by the acute pharmacological blockade of CB1 receptors, but inhibited by the CB1 agonist WIN55,212-2. CONCLUSION The endogenous cannabinoid system is involved in the control of neuroplasticity as part of pain processing . Cannabinoids prevent the formation of LTP in the amygdala via activation of CB1 receptors. Synoptic transmission and the inducibility of LTP were not influenced by the acute pharmacological blockade of CB1 receptors, but inhibited by the CB1 agonist Win55,212-2.

Flupirtine is a centrally acting nonopioid analgesic with muscle-relaxant properties. Flupirtine has been found to activate inwardly rectifying potassium conductances and hence to indirectly inhibit the activation of NMDA receptors. NMDA receptor activation is crucial for the induction of long-term potentiation (LTP) of synaptic transmission, which is considered as cellular correlate of learning and memory and of central sensitization in chronic pain states. Although flupirtine has been widely used for the management of pain, its effects on synaptic plasticity have not yet been investigated. We, therefore, performed extracellular and whole-cell patch-clamp recordings in hippocampal slices of mice to examine the effects of flupirtine on synaptic plasticity and neuronal membrane properties. Excitatory postsynaptic potentials (EPSPs) in the CA1 region were evoked alternately by stimulating two independent Schaffer collateral-commissural inputs. LTP and long-term depression (LTD) were induced by different stimulation paradigms (100 Hz, 10 Hz, 5 Hz, and 1 Hz). Flupirtine (30 microM) diminished the degree of LTP and enhanced LTD. This effect is most likely due to the hyperpolarization of CA1 pyramidal neurons and the reduction of their input resistance found after application of flupirtine. The observed effects on synaptic strength could underly the beneficial effects of flupirtine on different types of chronic pain.

Attempts have been made to attribute the particular features of general anaesthesia such as hypnosis, analgesia, amnesia and autonomic stability to certain brain regions. In the present study, we examined the effects of the commonplace volatile anaesthetic isoflurane on synaptic transmission in an in vitro slice preparation of the murine amygdala. Despite the established role of this limbic structure in the formation of aversive memories, conditioned fear and anxiety, as well as pain processing and regulation of sympathetic tone, the influence of volatile anaesthetics on synaptic signalling has not yet been investigated in this region of the brain. Evoked postsynaptic currents were monitored from principal neurons in the basolateral nucleus of the amygdala by means of patch-clamp recording. The mixed postsynaptic currents were mediated by non-NMDA, NMDA, GABA A and GABA B receptors. Isoflurane added to the perfusion medium reduced the strength of synaptic signalling following the activation of non-NMDA, NMDA, and GABA B receptors, whereas the GABA A receptor-mediated responses were enhanced. The overall reduction of neuronal excitability was also reflected in a reduction of field potential amplitudes. Isoflurane neither changed the membrane resting potential nor the input resistance of principal neurons in the amygdala. The present results may contribute to the understanding of how stress reactions and long-lasting neuroplastic processes are suppressed under general anaesthesia.

We examined whether sensory deprivation during formation of the cortical circuitry influences the pattern of intracortical single-cell connections in rat barrel cortex. Excitatory postsynaptic potentials (EPSPs) from layer 2/3 (L2/3) pyramidal neurons were recorded in vitro using patch-clamp techniques. In order to evoke EPSPs, presynaptic neurons were stimulated by photolytically applied glutamate, thus generating action potentials. Synaptic connections between the stimulated and the recorded neuron were identified by the occurrence of PSPs following photostimulation. Sensory deprivation changed the pattern of projections from L4 and L2/3 neurons to L2/3 pyramidal cells. In slices of non-deprived rats 86% of the total presynaptic neurons were located in the first and only 10% in the second barrel column. Deprivation changed these values to 67% and 26%, respectively. Therefore, the probability of presynaptic cells projecting to L2/3 neurons was shifted from adjacent to more remote barrel columns. These results indicate that deprivation of sensory input influences the pattern of intracortical connections.

The minimum alveolar concentration (MAC) of a volatile anesthetic defines anesthetic potency in terms of a suppressed motor response to a noxious stimulus. Therefore, the MAC of an anesthetic might in part reflect depression of motor neuron excitability. In the present study we evaluated the effect of isoflurane (ISO) on neurons in the substantia gelatinosa driven synaptically by putative nociceptive inputs in an in vitro spinal cord preparation of the rat. Whole-cell patch-clamp recordings were performed in neurons with their soma in the substantia gelatinosa of transverse rat spinal cord slices. We investigated the effect of ISO on excitatory postsynaptic currents (EPSC) evoked by dorsal root stimulation (eEPSC), spontaneous (sEPSC), and miniature (mEPSC) EPSC. ISO reversibly reduced the amplitude of eEPSC to 39% +/- 22% versus control. ISO decreased the frequency of sEPSC and mEPSC to 39% +/- 26% and 63% +/- 7%. Neither the amplitudes nor the kinetics of mEPSC and sEPSC were altered by ISO. We conclude that ISO depresses glutamatergic synaptic transmission of putative nociceptive primary-afferent inputs, presumably by reducing the release of the excitatory transmitter. This effect may contribute to an antinociceptive action of volatile anesthetics at the spinal cord level.The present electrophysiological in vitro experiments provide evidence that the volatile anesthetic isoflurane reduces excitatory transmitter release at the first site of synaptic integration of nociceptive inputs, the spinal cord superficial dorsal horn. This effect may contribute to the anesthetic action of volatile anesthetics at the spinal cord level.

The serotonin type 3 (5-HT(3)) receptor is the only ligand-gated ion channel receptor for serotonin (5-HT). 5-HT(3) receptors play an important role in modulating the inhibitory action of dopamine in mesocorticolimbic brain regions. Neuroleptic drugs are commonly thought to exert their psychopharmacological action mainly through dopamine and serotonin type 2 (5-HT(2)) receptors. Except for clozapine, a direct pharmacological interaction of neuroleptics with 5-HT(3) receptors has not yet been described. Using the concentration-clamp technique, we investigated the effects of flupentixol, various phenothiazines, haloperidol, clozapine and risperidone on Na(+)-inward currents through 5-HT(3) receptors stably expressed in human embryonic kidney 293 cells, and through endogenous 5-HT(3) receptors of murine N1E-115 neuroblastoma cells. In addition, we studied their effects on Ca(2+) influx, measured as a change in intracellular Ca(2+) concentrations ([Ca(2+)](i)). All neuroleptic drugs, but not risperidone, antagonized Na(+)- and Ca(2+)-inward currents evoked by 5-HT (10 microM for 2 s and 1 microM, respectively) in a voltage-independent manner. Only clozapine was a competitive antagonist, while all other compounds turned out to be noncompetitive. Fluphenazine and haloperidol affected membrane anisotropy at concentrations below their IC(50) values, indicating that a change in membrane anisotropy might contribute to their antagonistic effect at the 5-HT(3) receptor. Only structure analogues of flupentixol and fluphenazine with a lipophilic side chain were potent antagonists against 5-HT-evoked Na(+) and Ca(2+) currents. Since 5-HT(3) receptors modulate mesolimbic and mesocortical dopaminergic activity, the functional antagonism of neuroleptics at 5-HT(3) receptors may contribute to their antipsychotic efficacy and may constitute a not yet recognized pharmacological principle of these drugs.

Abnormally high spiking activity can damage neurons. Signaling systems to protect neurons from the consequences of abnormal discharge activity have been postulated. We generated conditional mutant mice that lack expression of the cannabinoid receptor type 1 in principal forebrain neurons but not in adjacent inhibitory interneurons. In mutant mice,the excitotoxin kainic acid (KA) induced excessive seizures in vivo. The threshold to KA-induced neuronal excitation in vitro was severely reduced in hippocampal pyramidal neurons of mutants. KA administration rapidly raised hippocampal levels of anandamide and induced protective mechanisms in wild-type principal hippocampal neurons. These protective mechanisms could not be triggered in mutant mice. The endogenous cannabinoid system thus provides on-demand protection against acute excitotoxicity in central nervous system neurons.

The distribution of glutamate receptor subtypes on the surface of neurons is highly relevant for synaptic transmission and signal processing. In the present study we investigated the location and properties of functional kainate receptors (KARs) on the somatodendritic membrane of rat neocortical layer V pyramidal neurons. Infrared-guided laser stimulation was used to apply glutamate photolytically to the soma and various sites along the apical dendrite. Electrical currents, resulting from the activation of pharmacologically isolated KARs, were measured by whole-cell patch-clamp recording. In addition, KARs on somatic and dendritic outside-out patches were activated while still within the brain tissue. We found that functional KARs are located on the entire somatodendritic membrane that was examined. Fast kinetics, a linearI-Vrelationship, and a relatively high single-channel conductance are characteristic features of these receptors. We provide evidence that the unitary properties of somatic and dendritic KARs are identical. Regarding the subcellular distribution of KARs, our results indicate that the density of these receptors increases toward the distal dendrite. They are located mainly at extrasynaptic sites but also mediate fast synaptic signaling triggered by afferent stimulation. The differential distribution speaks in favor of a selective targeting of KARs on central neurons and may reflect a mechanism for a location-dependent regulation of synaptic efficacy. Furthermore, it is feasible to assume that extrasynaptic KARs could be activated by a “spillover” of synaptically released glutamate, ambient glutamate in the CSF, or glutamate released from adjacent astrocytes.

Activation of adrenoreceptors modulates synaptic transmission in the basolateral amygdala. Here, we investigated the effects of alpha2-adrenoreceptor activation on long-term depression and long-term potentiation in an in vitro slice preparation of the mouse basolateral amygdala. Field potentials and excitatory postsynaptic currents were evoked in the basolateral amygdala by stimulating the lateral amygdala. Norepinephrine (20 micro m) reduced synaptic transmission and completely blocked the induction of long-term potentiation and long-term depression. The alpha2-adrenoreceptor antagonist yohimbine (2 micro m) reversed this effect. The alpha2-adrenoreceptor agonist clonidine (10 micro m) mimicked the effects of norepinephrine. The Gi/o-protein inhibitor pertussis toxin (5 micro g/mL) reversed the effect of clonidine. Long-term depression was blocked in the presence of omega-conotoxin GVIA, but not omega-agatoxin IVA. Clonidine inhibited voltage-activated Ca2+ currents mediated via N- or P/Q-type Ca2+-channels. The inhibitory action of clonidine on long-term depression was reversed when inwardly rectifying K+-channels were blocked by Ba2+ (300 micro m). The present data suggest that alpha2-adrenoreceptor activation impairs the induction of long-term depression in the basolateral amygdala by a Gi/o-protein-mediated inhibition of presynaptic N-type Ca2+-channels and activation of inwardly-rectifying K+-channels.

Homomeric complexes of 5-HT(3A) receptor subunits form a ligand-gated ion channel. This assembly does not fully reproduce the biophysical and pharmacological properties of native 5-HT(3) receptors which might contain the recently cloned 5-HT(3B) receptor subunit. In the present study, heteromeric assemblies containing human 5-HT(3A) and 5-HT(3B) subunits were expressed in HEK 293 cells to detail the functional diversity of 5-HT(3) receptors. We designed patch-clamp experiments with homomeric (5-HT(3A)) and heteromeric (5-HT(3AB)) receptors to emphasize the kinetics of channel activation and desensitization. Co-expression of the 5-HT(3B) receptor subunit reduced the sensitivity for 5-HT (5-HT(3A) receptor: EC(50) 3 micro M, Hill coefficient 1.8; 5-HT(3AB) receptor: EC(50) 25 micro M, Hill coefficient 0.9) and markedly altered receptor desensitization. Kinetic modeling suggested that homomeric receptors, but not heteromeric receptors, desensitize via an agonist-induced open-channel block. Furthermore, heteromeric 5-HT(3AB) receptor assemblies recovered much faster from desensitization than homomeric 5-HT(3A) receptor assemblies. Unexpectedly, the specific 5-HT(3) receptor agonist mCPBG induced an open-channel block at both homomeric and heteromeric receptors. Because receptor desensitization and resensitization massively affect amplitude, duration, and frequency of synaptic signaling, these findings are evidence in favor of a pivotal role of subunit composition of 5-HT(3) receptors in serotonergic transmission.

The synaptic site of expression of long-term potentiation (LTP) and long-term depression (LTD) is still a matter of debate. To address the question of presynaptic versus postsynaptic expression of neocortical LTP and LTD in a direct approach, we measured the glutamate sensitivity of apical dendrites of layer 5 pyramidal neurons during LTP and LTD. We used infrared-guided laser stimulation to release glutamate from its “caged” form with high spatial and temporal resolution. Responses to photolytically released glutamate and synaptically evoked EPSPs were recorded with patch-clamp pipettes from the neuronal somata. LTP and LTD could be induced by electrical stimulation at the same synapses in succession. The NMDA receptor-dependent LTD was accompanied by a decrease in the dendritic glutamate sensitivity, suggesting a postsynaptic expression of neocortical LTD. In contrast, LTP was never accompanied by a change in the dendritic glutamate sensitivity. A possible explanation for this finding is a presynaptic expression of neocortical LTP. Another set of experiments corroborated these results: Photolytic application of glutamate with a frequency of 5 Hz caused a long-lasting Ca(2+) and NMDA receptor-dependent decrease in the dendritic glutamate sensitivity. In contrast, LTP of dendritic glutamate sensitivity was never induced by photostimulation, despite several experimental modifications to prevent washout of the induction mechanism and to induce a stronger postsynaptic Ca(2+) influx. In conclusion, our findings provide strong evidence for a postsynaptic expression of neocortical LTD and favor a primarily presynaptic locus of neocortical LTP.

Acquisition and storage of aversive memories is one of the basic principles of central nervous systems throughout the animal kingdom. In the absence of reinforcement, the resulting behavioural response will gradually diminish to be finally extinct. Despite the importance of extinction, its cellular mechanisms are largely unknown. The cannabinoid receptor 1 (CB1) and endocannabinoids are present in memory-related brain areas and modulate memory. Here we show that the endogenous cannabinoid system has a central function in extinction of aversive memories. CB1-deficient mice showed strongly impaired short-term and long-term extinction in auditory fear-conditioning tests, with unaffected memory acquisition and consolidation. Treatment of wild-type mice with the CB1 antagonist SR141716A mimicked the phenotype of CB1-deficient mice, revealing that CB1 is required at the moment of memory extinction. Consistently, tone presentation during extinction trials resulted in elevated levels of endocannabinoids in the basolateral amygdala complex, a region known to control extinction of aversive memories. In the basolateral amygdala, endocannabinoids and CB1 were crucially involved in long-term depression of GABA (gamma-aminobutyric acid)-mediated inhibitory currents. We propose that endocannabinoids facilitate extinction of aversive memories through their selective inhibitory effects on local inhibitory networks in the amygdala.

Abstinence from chronic morphine results in characteristic withdrawal symptoms in humans and experimental animals. Despite a large number of studies, the cellular mechanisms underlying opiate withdrawal symptoms are not clearly understood, in particular, the regulation of micro-opioid receptor function during this process. The present study investigated the micro-opioid receptor-stimulated G-protein activity using guanylyl 5'-[gamma-[35S]thio]-triphosphate ([35S]-GTPgammaS) autoradiography. [35S]-GTPgammaS binding was performed using coronal rat brain sections (20 microm) in the presence or absence of the micro-opioid selective agonist [D-Ala(2),N-MePhe(4)Gly(5)-ol] enkephalin (DAMGO). In experiment 1, rats (male, Sprague-Dawley) were injected every 12 h with increasing doses of morphine (5-100 mg/kg, s.c.) for 12 days; a separate group of rats which received saline injections served as control. Opiate withdrawal was induced by abstinence from morphine. Thirty-six hours after the last morphine injection, spontaneous withdrawal symptoms were assessed. Rats were then decapitated and brains rapidly removed. In experiment 2, withdrawal symptoms were precipitated with the opioid receptor antagonist naloxone (1 mg/kg). Brains were taken at 5, 10, 20 and 60 min after naloxone injection. In experiment 3, morphine dependence was induced by implantation of three morphine pellets (75 mg per pellet). After 7 days, withdrawal symptoms were precipitated by naloxone (1 mg/kg) and brains were removed 30 min after naloxone injection. [35S]-GTPgammaS binding was measured in the locus coeruleus, nucleus parabrachialis, nucleus accumbens and central nucleus of amygdala. Although clear withdrawal symptoms were observed in all morphine-withdrawn rats, no significant changes in [35S]-GTPgammaS binding were detected in animals undergoing withdrawal. The present lack of differences between morphine-withdrawn and control rats indicates that micro-opioid receptor-stimulated G-protein activity is not modulated by chronic morphine administration and is not involved in the expression of opiate withdrawal.

Recombinant α 1 β 2 γ 2L γ-aminobutyric acid (A) receptor (GABA A R) channels expressed in human embryonic kidney (HEK293) cells were used for patch-clamp experiments. The currents activated by brief pulses of GABA (10 −4 M) applied with a device for fast solution exchange to cells clamped in the whole-cell configuration mimicked GABA A R-mediated inhibitory postsynaptic currents. Isoflurane (ISO) at clinically relevant concentrations (0.6 mM) decreased the amplitude and prolonged the decay of the GABA-evoked response. To further detail the mechanism underlying the prolonged decay time, we made simulations based on these measurements. These simulations suggest that ISO slows the rate of GABA unbinding from the receptor. Under these conditions, ISO increases the GABA-induced charge transfer and, thus, could enhance GABAergic inhibition despite the concomitant open-channel block causing the decrease in the current amplitude.

The type 3 serotonin (5-HT(3)) receptor is a ligand-gated ion channel. In concentration-clamp experiments, we investigated the effects of the uncompetitive N-methyl-D-aspartate (NMDA) receptor antagonists memantine, amantadine and MRZ 2/579 on 5-HT receptors stabley expressed in HEK-293 cells and on native 5-HT(3) receptors in the N1E-115 cell line. All agents antagonized serotonin (10 microM)-induced inward currents with similar potency to that reported for NMDA receptors. This effect was characterized by inducing a pronounced receptor desensitization, and was probably non-competitive and voltage-independent. In contrast, (S)-ketamine was much weaker as an antagonist of 5-HT(3) receptors than NMDA receptors. Similar effects on 5-HT(3) receptors have been reported previously for a variety of anti-depressants and it is possible that the clinical anti-depressant effects reported for both memantine and amantadine are mediated, at least in part, by antagonistic effects at 5-HT(3) receptors.

The distribution of functional neurotransmitter receptors on the surface of neurons is highly relevant for synaptic transmission and signal processing. To map functional GABA(A) and GABA(B) receptors on the somadendritic membrane of rat neocortical layer V pyramidal neurons we used patch-clamp recording in combination with infrared-guided laser stimulation to release gamma-aminobutyric acid (GABA) photolytically. The data strongly suggest that relatively more GABA(A) receptors are located at the apical dendrite and relatively more GABA(B) receptors near the soma. Such a specific distribution of GABA(A) and GABA(B) receptors may serve to compensate for differences in electrotonic voltage attenuation between GABA(A) and GABA(B) receptor-mediated inhibitory postsynaptic potentials (IPSPs).

This study investigated whether the nitric oxide pathway was involved in the central effects of Delta(9)-tetrahydrocannabinol (Delta(9)-THC), the major psychoactive constituent of cannabis sativa. Body temperature, nociception and locomotion were measured in neuronal nitric oxide synthase (nNOS) knock-out (KO) mice and wild-type (WT) controls after intraperitoneal application of Delta(9)-THC. These Delta(9)-THC-induced effects are known to be mediated through the brain-type cannabinoid receptor 1 (CB1). Therefore, in situ hybridization (ISH) experiments were performed in the adult murine brain to determine possible changes in CB1 mRNA levels in nNOS-KO, compared with WT mice, and to reveal brain areas where CB1 and nNOS were coexpressed in the same neurons. We found that an intraperitoneal injection of 10 mg/kg Delta(9)-THC led to the same increase in the hot plate latencies in both genotypes, suggesting that Delta(9)-THC-mediated antinociception does not involve nNOS. In contrast, a significant Delta(9)-THC-induced decrease of body temperature and locomotor activity was only observed in WT, but not in nNOS-KO mice. ISH revealed significantly lower levels of CB1 mRNA in the ventromedial hypothalamus (VMH) and the caudate putamen (Cpu) of the nNOS-KO animals, compared with WT mice. Both areas are known to be among the regions involved in cannabinoid-induced thermoregulation and decrease of locomotion. A numerical evaluation of nNOS/CB1 coexpression showed that approximately half of the nNOS-positive cells in the dorsolateral Cpu also express low levels of CB1. ISH of adjacent serial sections with CB1 and nNOS, revealed expression of both transcripts in VMH, suggesting that numerous nNOS-positive cells of VMH coexpress CB1. Our findings indicate that the nitric oxide pathway is involved in some, but not all of the central effects of Delta(9)-THC.

Metabotropic glutamate receptors (mGluR) have been shown to play a role in the modulation of acute and inflammatory pain. Additionally, we have recently detected time-dependent changes in the mRNA expression of several mGluR subtypes in thalamic nuclei of monoarthritic (MA) rats. In the present study, mGluR1, -3, -4, and -7 subtype mRNA expression was analyzed by in situ hybridization with radioactively labelled oligonucleotide probes in cerebral cortical regions of normal and MA rats at 2, 4, and 14 days of the disease. The mGluR1, -4, and -7 mRNAs were at background level in normal rats and did not change in MA animals. In contrast, mGluR3 mRNA expression was abundant in normal rats and was significantly increased in cortical areas of MA rats at all time points. Higher changes were detected bilaterally at 4 days, predominantly in layers IV/V, in the motor, primary, and secondary somatosensory cortices (average increases of 50-75%), but maximum rises occurred in the contralateral cingulate cortex (+138%). No changes were detected in the auditory cortex. The present data show an up-regulation of mGluR3 mRNA expression in the motor, somatosensory, and limbic cortices of MA rats. This possibly reflects the occurrence of central mechanisms counteracting the increased transmission of nociceptive input arising from the inflamed paw and the impaired motor behavior of these rats. Changes in the cingulate cortex may be related to the motivational-affective component of nociception.

A reduction in glucocorticoid receptor (GR) function leads to hippocampus-dependent allocentric spatial learning deficits, altered novelty exploration and disrupted hippocampal long-term potentiation (LTP) in transgenic mice expressing a GR antisense construct. After continuous long-term treatment of these mice with moclobemide (a reversible inhibitor of monoamine oxidase A), spatial navigation performance but not accuracy improved during initial acquisition. These changes were associated with a shift of the threshold for the induction of hippocampal LTP at low stimulation frequencies. Moreover, novel object exploration increased in both control and transgenic animals following long-term treatment with moclobemide. These findings open the possibility that antidepressants might improve hippocampal function under conditions of impaired stress hormone regulation, and that these drugs might in part act through this mechanism to attenuate cognitive deficiency in disorders such as depression.

We investigated the interactions between recombinant gamma-aminobutyric acid receptor complex (GABA(A)R) and nitrous oxide (N(2)O) or xenon (Xe). Human embryonic kidney cells (HEK 293) were transfected with rat cDNA for alpha(1)beta(2)gamma(2L) or for alpha(1)beta(2) recombinant GABA(A)R subunits. Patch clamp techniques were used in the whole-cell mode to evaluate the effect of N(2)O and Xe on GABA-induced currents. A piezo-driven "liquid filament switch" was used for fast application. Both N(2)O (100%, 29.2 mM) and Xe (100%, 3.9 mM) reversibly increased GABA-induced currents through the alpha(1)ss(2)gamma(2L) and the alpha(1)beta(2) GABA(A)R channels. The potentiating effect of N(2)O or Xe on peak currents was prominent at small GABA concentrations (10(-7) to 10(-5) M). The addition of N(2)O or Xe increased the efficacy of GABA (10(-7) to 10(-3) M). Both N(2)O and Xe significantly decreased the risetime((10%-90%)) of the currents elicited by small GABA concentrations. At the concentrations used, neither N(2)O nor Xe had an intrinsic effect. We conclude that, similar to other anesthetics, both N(2)O and Xe increase the efficacy of GABA at the GABA(A)R and enhance inhibitory GABAergic synaptic transmission.

Electrophysiological and behavioural experiments were performed in transgenic mice expressing a dominant-negative form of cAMP response element-binding protein (CREBA133) in the limbic system. In control littermate in vitro slice preparation, tetanizing the lateral amygdala-basolateral amygdala (BLA) pathway with a single train (100 Hz for 1 s) produced short-term potentiation (STP) in the BLA. Five trains (10-s interstimulus interval) induced long-term potentiation (LTP), which was completely blocked by the N-methyl-D-aspartate (NMDA) receptor antagonist D(-)-2-amino-5-phosphonopentanoic acid (AP5; 50 microM). When GABAergic (gamma-aminobutyric acid) inhibition was blocked by picrotoxin (10 microM), LTP became more pronounced. Low-frequency stimulation (1 Hz for 15 min) induced either long-term depression (LTD) or depotentiation. LTD remained unaffected by AP5 (50 microM) or by the L- and T-type Ca2+-channel blockers nifedipine (20 microM) and Ni2+ (50 microM), but was prevented by picrotoxin (10 microM), indicating a GABAergic link in the expression of LTD in the BLA. When conditioned fear was tested, a mild impairment was seen in one of three transgenic lines only. Although high levels of mRNA encoding CREBA133 lead to downregulation of endogenous CREB, expression of LTP and depotentiation were unaltered in BLA of these transgenic animals. These results could suggest that residual CREB activity was still present or that CREB per se is dispensable. Alternatively, other CREB-like proteins were able to compensate for impaired CREB function.

The expression of metabotropic glutamate receptor (mGluR) mRNAs in the human cerebellar neocortex was investigated using in situ hybridization. mGluR1, 3 and 4 mRNAs were most abundant and widely distributed, whereas mGluR2, 5 and 7 mRNA expression was circumscript. mRNAs coding for mGluR1, 3, 4 and 7 were expressed in Purkinje cells. mGluR1 and 4 mRNAs detectable in granule cells and mGluR1, 2, 3, 4 and 7 mRNAs in Golgi cells. mGluR5 mRNA was detectable in putative Bergmann glia as well as mGluR3 mRNA, which was widely expressed in glial cells. mGluR8 mRNA expression was very low in a subset of stellate/basket cells of the molecular layer whereas mGluR6 mRNA was not detectable in the cerebellar cortex.

Following chronic alcohol treatment alterations in N-methyl- d -aspartate receptor subunit 1 and 2 (NR1 and NR2), mRNA and protein levels have been reported. The NR1 gene undergoes alternative RNA splicing, resulting in eight splice variants, which were shown to differ in their sensitivity to alcohol. Here, we studied mRNA and protein levels of NR1 splice variants in alcohol-preferring (AA) and alcohol-nonpreferring (ANA) rat lines under basal conditions (alcohol-naive), and following chronic alcohol consumption. mRNA levels of three NR1 splice variants (NR1-1, NR1-2, NR1-4), and the protein levels of NR1 (NR1-1/NR1-2), and of NR1 alternative C-terminus (NR1-3/NR1-4) were determined in the hippocampus and nucleus accumbens by competitive RT-PCR and Western blot analysis, respectively. No significant differences in NR1 mRNA, or protein levels were found in the nucleus accumbens between the two rat lines under basal conditions, or following chronic alcohol consumption. In the hippocampus of alcohol-naive rats, the NR1-4 mRNA content was significantly higher in ANA compared to AA rats, however, no significant difference could be detected at the protein level. Following chronic alcohol consumption, the protein level of the NR1 alternative C-terminus (NR1-3/NR1-4) was significantly higher in AA rats compared to the corresponding control. Taken together, these results suggest: (i) brain site-specific alterations in NMDA receptor subunit composition occur following chronic alcohol consumption. (ii) In the hippocampus, NR1 splice variant mRNA levels differ between AA and ANA rats. (iii) The mRNA levels and protein levels of NR1 splice variants are differentially affected by chronic alcohol consumption.

In a direct approach to elucidate the origin of long-term depression (LTD), glutamate was applied onto dendrites of neurons in rat neocortical slices. An infrared-guided laser stimulation was used to release glutamate from caged glutamate in the focal spot of an ultraviolet laser. A burst of light flashes caused an LTD-like depression of glutamate receptor responses, which was highly confined to the region of “tetanic” stimulation (

The development of chronic pain is associated with activity-dependent plastic changes in neuronal structures in the peripheral and central nervous system. In order to investigate the time-dependent processing of afferent noxious stimuli in the spinal cord we employed the quantitative autoradiographic 2-deoxyglucose technique in a model of chronic monoarthritic pain in the rat. Spinal metabolic activity was determined at various time-points (two, four and 14 days) after the injection of complete Freund's adjuvant into the left tibiotarsal joint. In addition, the effect of acute noxious mechanical stimulation of the arthritic joint was investigated at 14 days of monoarthritis. Local glucose utilization was determined in lumbar segments L2-L5, ipsi- and contralateral to the inflamed hindpaw, and compared with saline-injected controls. In general, monoarthritic animals had bilaterally increased metabolic activity in all laminae of the spinal cord. Detailing the time-course showed that in rats with two days of monoarthritis metabolic activity was significantly increased to a similar extent on both sides of all spinal laminae. In contrast, at four days, glucose utilization in deep laminae of the dorsal horn (laminae V-VI), the central gray area (laminae X) and the ventral horn (laminae VII-IX) tended to return to control levels. At 14 days of monoarthritis, however, metabolic activity showed a further increase in all laminae of the spinal cord. This increase was more pronounced on the side ipsilateral to inflammation, reaching 65% above corresponding control levels in laminae V, VI. Animals with 14 days of monoarthritis which were subjected to mechanical noxious stimulation of the arthritic joint displayed clear behavioral signs of acute pain. Although in this group metabolic activity was above control levels, it was lower than in animals with 14 days of monoarthritis that were not additionally stimulated. The data show not only a general increase of spinal cord metabolic activity during the time-course of the development of a chronic pain state, but also show a region-specific non-linear time profile. This may reflect the complexity of transducing and suppressive transmitter systems involved in the central processing of ongoing pain.