The novel antipsychotic cariprazine stabilizes gamma oscillations in rat hippocampal slices

Background and purpose: Gamma oscillations are fast rhythmic fluctuations of neuronal network activity ranging from 30 to 90 Hz that establish a precise temporal background for cognitive processes such as perception, sensory processing, learning, and memory. Alterations of gamma oscillations have been observed in schizophrenia and are suggested to play crucial roles in the generation of positive, negative, and cognitive symptoms of the disease. Experimental approach: In this study, we investigated the effects of the novel antipsychotic cariprazine, a D3 -preferring dopamine D3 /D2 receptor partial agonist, on cholinergically induced gamma oscillations in rat hippocampal slices from treatment-naïve and MK-801-treated rats, a model of acute first-episode schizophrenia. Key results: The D3 receptor-preferring agonist pramipexole effectively decreased the power of gamma oscillations, while the D3 receptor antagonist SB-277011 had no effect. In treatment-naïve animals, cariprazine did not modulate strong gamma oscillations but slightly improved the periodicity of non-saturated gamma activity. Cariprazine showed a clear partial agonistic profile at D3 receptors at the network level by potentiating the inhibitory effects when the D3 receptor tone was low and antagonizing the effects when the tone was high. In hippocampal slices of MK-801-treated rats, cariprazine allowed stabilization of the aberrant increase in gamma oscillation power and potentiated resynchronization of the oscillations. Conclusion and implications: Data from this study indicate that cariprazine stabilizes pathological hippocampal gamma oscillations, presumably by its partial agonistic profile. The results demonstrate in vitro gamma oscillations as predictive biomarkers to study the effects of antipsychotics preclinically at the network level

Capsaicin-Induced Changes in LTP in the Lateral Amygdala Are Mediated by TRPV1

The transient receptor potential vanilloid type 1 (TRPV1) channel is a well recognized polymodal signal detector that is activated by painful stimuli such as capsaicin. Here, we show that TRPV1 is expressed in the lateral nucleus of the amygdala (LA). Despite the fact that the central amygdala displays the highest neuronal density, the highest density of TRPV1 labeled neurons was found within the nuclei of the basolateral complex of the amygdala. Capsaicin specifically changed the magnitude of long-term potentiation (LTP) in the LA in brain slices of mice depending on the anesthetic (ether, isoflurane) used before euthanasia. After ether anesthesia, capsaicin had a suppressive effect on LA-LTP both in patch clamp and in extracellular recordings. The capsaicin-induced reduction of LTP was completely blocked by the nitric oxide synthase (NOS) inhibitor L-NAME and was absent in neuronal NOS as well as in TRPV1 deficient mice. The specific antagonist of cannabinoid receptor type 1 (CB1), AM 251, was also able to reduce the inhibitory effect of capsaicin on LA-LTP, suggesting that stimulation of TRPV1 provokes the generation of anandamide in the brain which seems to inhibit NO synthesis. After isoflurane anesthesia before euthanasia capsaicin caused a TRPV1-mediated increase in the magnitude of LA-LTP. Therefore, our results also indicate that the appropriate choice of the anesthetics used is an important consideration when brain plasticity and the action of endovanilloids will be evaluated. In summary, our results demonstrate that TRPV1 may be involved in the amygdala control of learning mechanisms

The effect of TRPV1 receptors on the synaptic plasticity in the lateral nucleus of the amygdala

Bei dieser Arbeit stand die Wirkung des TRPV1-Rezeptors auf erregende Projektionsneuronen im Mittelpunkt. Es sollte geklärt werden, wie sie die synaptische Plastizität modulieren können und ob Sensibilisatoren die Ergebnisse beeinflussen. Im Rahmen dieser Doktorarbeit konnte gezeigt werden, dass der TRPV1-Rezeptor hauptsächlich an der präsynaptischen Membran von Projektionsneuronen lokalisiert ist. Capsaicin hat eine gesteigerte Glutamatfreisetzung bewirkt, wodurch die Kurzzeit-plastizität beeinflusst sein kann. Die TRPV1-Rezeptoraktivierung hat einen signifikanten, hemmenden Einfluss auf die Stärke einer LTP. In TRPV1-knockout-Mäusen ist die Stärke der LTP ebenfalls signifikant reduziert. Innerhalb dieser Arbeit konnte erstmalig bewiesen werden, dass durch eine Hoch-frequenzstimulation die Synthese von Stickstoffmonoxid in Gehirnschnitten des lateralen Kerns der Amygdala erhöht wird. Es ist zu beachten, dass die NO-Produktion auch schon vor einer HFS konstant stattfindet. Es konnte eine Sensibilisierung des TRPV1-Rezeptors durch Isofluran, Ethanol und Serotonin nachgewiesen werden. Ethanol und Serotonin wirken supprimierend auf die LA-LTP. Bei Koapplikation mit Capsaicin ist die LA-LTP erhöht. Bei Narkosen sollte eine Isoflurankonzentrationen von unter 4 % verwendet werden, um die Messergebnisse nicht zu beeinflussen. Die durch Serotonin verursachte Sensibilisierung des TRPV1-Rezeptors wird möglicherweise durch den 5-HT2B- und/oder 5-HT2C-Rezeptor vermittelt. Im Rahmen dieser Dissertation konnte erstmalig gezeigt werden, dass der Forced Swimming Test einen suppressiven Effekt auf die LA-LTP hat. Dies unterstützt andere Stressstudien von Amygdala und Hippokampus. Es konnte gezeigt werden, dass die Akti-vierung des Rezeptors diesen stressinduzierten, suppressiven Effekten entgegen wirkt. Da der TRPV1-Rezeptor ein mögliches Ziel bei der Behandlung von Angststörungen ist, ist es notwendig dessen Wirkungsweise zu kennen, um geeignete Pharmaka für die Behandlung von Patientinnen und Patienten herstellen zu können. Weiterhin könnte eine gezielte pharmakologische Beeinflussung von TRPV1-Ionenkanälen eine praktikable Methode zur Behandlung von durch Stress verursachten, krankhaften Hyperaktivitäten der Amygdala mit daraus resultierenden Verhaltensstörungen sein.The focus of this work was on the effect of the TRPV1 receptors at glutamatergic excitatory projection neurons. It was of interest how their activation modulates the synaptic plasticity and how possible sensitizers would change the outcome. In the framework of this doctoral thesis it could be shown that the TRPV1 receptors are mainly localized at the presynaptic membrane of projection neurons. This receptor type can modulate the short term plasticity. TRPV1 activation leads to an increased release of glutamate. The strength of the LTP is significantly influenced when these receptor types are activated. Since the strength of LTP is reduced in TRPV1 knockout mice the influence without activation is significant as well. Within this work the first evidence could be reported that high frequency stimulation leads to an increase in the production of nitric oxide in horizontal brain slices of the lateral nucleus of the amygdala. It is to recognize that NO is produced continuously before the HFS as well. TRPV1 activation increases the NO production. It could be shown that the TRPV1 receptor can be sensitized by isoflurane, ethanol, and serotonin. Ethanol and serotonin cause a reduction of LA-LTP. In co application with capsaicin they increase LA-LTP. To not influence the experimental results isoflurane should be used in a concentration of 4 % or less for anaesthesia. The sensitization of the TRPV1 receptor caused by serotonin is presumably mediated by the 5-HT2B, the 5-HT2C receptor, or both. In the context of this dissertation it could be shown for the first time that the forced swimming test has a suppressive effect on the LTP in the LA. This finding supports other stress studies in the amygdala and the hippocampus. Further it can be reported that an activation of the TRPV1 receptor can counteract this stress induced suppressive effect. Since the TRPV1 receptor is a possible target in the treatment of fear disorders it is essential to know its way of action in order to develop suitable pharmaceutics for the treatment of patients. Additionally, pharmacological manipulation of the TRPV1 receptor could be a method for treating stress induced pathological hyperactivities in the amygdala with its resulting behaviour disorders

Properties of spontaneous PSCs in the absence and presence of TTX.

<p>(A) Continuous recording shows spontaneous EPSCs in the presence of TTX (mEPSCs) from a LA projection neuron under control conditions and 10 min after bath application of capsaicin (1 µM), V_m = −70 mV. (B) Continuous recording shows spontaneous IPSCs in the presence of TTX (mIPSCs) from another LA projection neuron under control conditions and 10 min after bath application of capsaicin (1 µM), V_m = 0 mV. (C) Summary data shows mean frequency of mEPSCs, mIPSCs and sIPSCs under control conditions and in the presence of capsaicin, SEM is indicated by error bars. (D) Amplitude distributions of mIPSCs recorded in under control conditions (black) and in the presence of 1 µM capsaicin (grey). Superimposed lines show cumulative amplitude histograms (p>0.05, Kolmogoroff-Smirnov test). (E) Continuous recording shows spontaneous IPSCs in the absence of TTX (sIPSCs) under control conditions and 10 min after bath application of capsaicin (1 µM), V_m = −70 mV. (F) Amplitude distributions of sIPSCs recorded in under control conditions (black) and in the presence of 1 µM capsaicin (grey). Superimposed lines show cumulative amplitude histograms.</p

Isoflurane anesthesia before euthanasia instead of ether caused a capsaicin-induced enhancement of LA-LTP in horizontal slices derived from adult mice.

<p>(A) HFS-induced LTP is increased in magnitude by 1 µM capsaicin in comparison with control. This increase can be blocked by the specific TRPV1 antagonist AMG9810. (B) Capsaicin-induced LTP enhancement is absent in TRPV1^−/− mice. Representative traces were recorded 5 min prior to tetanus (dashed lines) and 60 min after tetanus (solid lines). (C) Bar histogram of data points averaged 57 to 60 min after HFS and normalized with respect to baseline (mean ± SEM). Significant differences are indicated. *<i>p</i>≤0.05.</p

TRPV1 protein expression within the amygdala.

<p>(A–C) These graphs illustrate the densities of cell nuclei (visualized with DAPI), neurons (visualized with NeuroTrace), cells expressing TRPV1 (visualized by immunohistochemistry) as well as the densities of neuronal and non-neuronal cells expressing TRPV1 in the lateral nucleus (A), basolateral (B) and central nucleus (C) of the amygdala. (D) The numerical neuronal density of TRPV1 immunopositive neurons in the basolateral amygdala (LA, BL) is significantly higher than in the CE, as determined by ANOVA followed by a Tukey's posthoc test (ns: non-significant; **: p≤0.01; ***: p≤0.001). (E–G) Example of a triple-stained section (DAPI, NeuroTrace and anti-TRPV1; lateral nucleus of the amygdala). NeuroTrace was used to visualize neuronal cells (in green, E). In the same section, binding of TRPV1 antibodies was visualized with Cy5 (in red, F). Figure G displays the merged images of E and F (in each case DAPI (in blue) was used to visualize cell nuclei).</p

The capsaicin-induced reduction of LTP recorded in horizontal slices is blocked by TRPV1 antagonists and is absent in TRPV1 deficient mice.

<p>(A) Input-output curves as evoked by single stimuli applied at EC fibers. Drug-free control recordings (n = 15) did not differ from recordings made in capsaicin-treated slices (1 µM cap: n = 15; 10 µM cap: n = 8). (B) EC stimulation caused a stable LA-LTP in horizontal slices derived from adult male mice. Bath-applied capsaicin (cap) induced a dose-dependent reduction in the magnitude of LA-LTP. Data points represent averaged amplitudes (mean ± SEM) normalized with respect to baseline values. Representative traces were recorded 5 min prior to tetanus (dashed lines) and 60 min after tetanus (solid lines). (C) Capsazepine completely blocks the capsaicin-induced reduction of LTP only at a concentration of 50 µM. (D) The capsaicin-induced reduction of LA-LTP could be also blocked by the specific TRPV1 antagonist AMG9810. (E) Bar histogram of data points averaged 57 to 60 min after HFS and normalized with respect to baseline (mean ± SEM). Significant differences are indicated. *<i>p</i><0.05. (F) In TRPV1^−/− mice the inhibitory effect of 1 µM capsaicin was absent. HFS: high-frequency stimulation.</p

EC-induced LTP is also suppressed by capsaicin in coronal brain slices.

<p>This suppression cannot be blocked in the presence of 10 µM SR95531. (A) In extracellular recordings the magnitude of LA-LTP obtained in coronal slices derived from juvenile mice is lower than that in horizontal slices. Nevertheless, capsaicin reduced LA-LTP in both instances. (B) In patch clamp recordings the complete blockade of GABA_A receptors did not change the inhibitory effect of capsaicin on LA-LTP obtained in coronal slices derived from adult mice. Representative traces were recorded 5 min prior to tetanus (dashed lines) and 60 min and 30 min, respectively, after tetanus (solid lines) in the lateral nucleus of the amygdala.</p