Endogén ligandok interakciója gerincvelői szinten = Interaction of endogenous ligands at the spinal level

Az OTKA pályázat keretében több irányban végeztünk kutatásokat a fájdalom mechanizmusok tisztázása érdekében. Meghatároztuk számos, a cannabinoid receptoron ható endogén ligand fájdalomcsillapító hatását és interakciójukat gerincvelői szinten. Az endomorphin-1, agmatin, adenozin, kinurénsav hármas és négyes kombinációban adva kedvező interakciót mutattak, így kombinációban tovább csökkenthető az alkalmazott dózis, s ezzel a mellékhatások veszélye tovább mérsékelhető. Kísérleteket végezünk a viszketésben szerepet játszó spinális mechanizmusok feltárására is. Az intratekálisan adott naltrexon csökkentette a viszketési magatartást, az NMDA antagonista ketamin nem befolyásolta azt jelentős mértékben, ugyanakkor az endogén kinurénsav már hatékony volt. Kifejlesztettünk egy szubkrónikus izületi gyulladásos modellt, és meghatároztuk számos, az izületbe injektált endogén ligandok hatását az izületi gyulladás okozta mechanikus allodyniára. Laboratóriumunkban kifejlesztettünk egy komplex, krónikus szkizofrénia modellt, melyben az állatok hőfájdalom szenzitivitása megváltozott. A modell további jellemzésére egy Minimitter készüléket szereztünk be, mely lehetővé teszi a hőszabályozás és a motoros aktivitás folyamatos regisztrálását telemetriás módszerrel. Legújabb vizsgálatainkban a fiatalkori capsaicin deszenzitizáció fájdalomküszöböt befolyásoló hatását jellemeztük igazolva, hogy a deszenzitizáció elsősorban a gyulladásos fájdalomérzékenységet csökkenti. | Thanks to the OTKA grant we performed several experiments to characterize the pain mechanisms in different aspects. We determined the antinociceptive effects and the interactions of several endogenous cannabinoid ligands at spinal level. We found that the triple and quadruple combinations of endomorphin-1, agmatine, adenosin and kynurenic acid induced effective antinociception without any side effects. We investigated the spinal mechanisms responsible for pruritus. The intrathecally administered naltrexone and kynurenic acid significantly decreased the pruritic behavior, while ketamine did not influence it. A subchronic inflammatory model was developed and characterized. We showed the antinociceptive potency and interactions of several endogenous ligands at peripheral level. It is well-known that the schizophrenic patients show decreased pain sensitivity. We developed and characterized a new, complex, chronic schizophrenia model. We proved that these animals show increased pain threshold. We invested a telemetric instrument (Minimitter) to further characterize this model (thermoregulation, motor activity). We also investigated the pain threshold changes in different pain models after juvenile capsaicin desensitization. We have shown that the inflammatory pain sensitivity decreased and the morphine has higher potency in these animals

Wisket rat model of schizophrenia : Impaired motivation and, altered brain structure, but no anhedonia

It is well-known that the poor cognition in schizophrenia is strongly linked to negative symptoms, including motivational deficit, which due to, at least partially, anhedonia. The goal of this study was to explore whether the schizophrenia-like Wisket animals with impaired motivation (obtained in the reward-based hole-board test), also show decreased hedonic behavior (investigated with the sucrose preference test). While neurochemical alterations of different neurotransmitter systems have been detected in the Wisket rats, no research has been performed on structural changes. Therefore, our additional aim was to reveal potential neuroanatomical and structural alterations in different brain regions in these rats. The rats showed decreased general motor activity (locomotion, rearing and exploration) and impaired task performance in the hole-board test compared to the controls, whereas no significant difference was observed in the sucrose preference test between the groups. The Wisket rats exhibited a significant decrease in the frontal cortical thickness and the hippocampal area, and moderate increases in the lateral ventricles and cell disarray in the CA3 subfield of hippocampus. To our knowledge, this is the first study to investigate the hedonic behavior and neuroanatomical alterations in a multi-hit animal model of schizophrenia. The results obtained in the sucrose preference test suggest that anhedonic behavior might not be involved in the impaired motivation obtained in the hole-board test. The neuropathological changes agree with findings obtained in patients with schizophrenia, which refine the high face validity of the Wisket model

Automating, Analyzing and Improving Pupillometry with Machine Learning Algorithms

The investigation of the pupillary light reflex (PLR) is a well-known method to provide information about the functionality of the autonomic nervous system. Pupillometry, a non-invasive technique, was applied to study the PLR alterations in a new, schizophrenia-like rat substrain, named WISKET. The pupil responses to light impulses were recorded with an infrared camera; the videos were automatically processed and features were extracted from the pupillograms. Besides the classical statistical analysis (ANOVA), feature selection and classification were applied to reveal the significant differences in the PLR parameters between the control and WISKET animals. Based on these results, the disadvantages of this method were analyzed and the measurement setup was redesigned and improved. The pupil segmentation method has also been adapted to the new videos. 2564 images were annotated manually and used to train a fully-convolutional neural network to produce pupil mask images. The method was evaluated on 329 test images and achieved 4% median relative error. With the new setup, the pupil detection became reliable and the new data acquisition offers robustness to the experiments

Caffeine-Induced Acute and Delayed Responses in Cerebral Metabolism of Control and Schizophrenia-like Wisket Rats

Recently, morphological impairments have been detected in the brain of a triple-hit rat schizophrenia model (Wisket), and delayed depressive effects of caffeine treatment in both control and Wisket animals have also been shown. The aims of this study were to determine the basal and caffeine-induced acute (30 min) and delayed (24 h) changes in the cerebral (18)fluorodeoxyglucose (F-18-FDG) uptake by positron emission tomography (PET) in control and Wisket rats. No significant differences were identified in the basal whole-brain metabolism between the two groups, and the metabolism was not modified acutely by a single intraperitoneal caffeine (20 mg/kg) injection in either group. However, one day after caffeine administration, significantly enhanced F-18-FDG uptake was detected in the whole brain and the investigated areas (hippocampus, striatum, thalamus, and hypothalamus) in the control group. Although the Wisket animals showed only moderate enhancements in the F-18-FDG uptake, significantly lower brain metabolism was observed in this group than in the caffeine-treated control group. This study highlights that the basal brain metabolism of Wisket animals was similar to control rats, and that was not influenced acutely by single caffeine treatment at the whole-brain level. Nevertheless, the distinct delayed responsiveness to this psychostimulant in Wisket model rats suggests impaired control of the cerebral metabolism

Ligand-Specific Regulation of the Endogenous Mu-Opioid Receptor by Chronic Treatment with Mu-Opioid Peptide Agonist

Since the discovery of the endomorphins (EM), the postulated endogenous peptide agonists of the mu-opioid receptors, several analogues have been synthesized to improve their binding and pharmacological profiles. We have shown previously that a new analogue, cis-1S,2R-aminocyclohexanecarboxylic acid2-endomorphin-2 (ACHC-EM2), had elevated mu-receptor affinity, selectivity, and proteolytic stability over the parent compound. In the present work, we have studied its antinociceptive effects and receptor regulatory processes. ACHC-EM2 displayed a somewhat higher (60%) acute antinociceptive response than the parent peptide, EM2 (45%), which peaked at 10 min after intracerebroventricular (icv) administration in the rat tail-flick test. Analgesic tolerance developed to the antinociceptive effect of ACHC-EM2 upon its repeated icv injection that was complete by a 10-day treatment. This was accompanied by attenuated coupling of mu-sites to G-proteins in subcellular fractions of rat brain. Also, the density of mu-receptors was upregulated by about 40% in the light membrane fraction, with no detectable changes in surface binding. Distinct receptor regulatory processes were noted in subcellular fractions of rat brains made tolerant by the prototypic full mu-agonist peptide, DAMGO, and its chloromethyl ketone derivative, DAMCK. These results are discussed in light of the recently discovered phenomenon, that is, the “so-called biased agonism” or “functional selectivity

Neurobehavioral impairments in ciprofloxacin-treated osteoarthritic adult rats

Background and purpose – Ciprofloxacin (CIP) is a broad-spectrum antibiotic widely used in clinical practice to treat musculoskeletal infections. Fluoroquinoloneinduced neurotoxic adverse events have been reported in a few case reports, all the preclinical studies on its neuropsychiatric side effects involved only healthy animals. This study firstly investigated the behavioral effects of CIP in an osteoarthritis rat model with joint destruction and pain, which can simulate inflammation-associated musculoskeletal pain. Furthermore, effects of CIP on regional brain-derived neurotrophic factor (BDNF) expression were examined given its major contributions to the neuromodulation and plasticity underlying behavior and cognition. Methods – Fourteen days after induction of chronic osteoarthritis, animals were administered vehicle, 33 mg/kg or 100 mg/kg CIP for five days intraperitoneally. Motor activity, behavioral motivation, and psychomotor learning were examined in a reward-based behavioral test (Ambitus) on Day 4 and sensorimotor gating by the prepulse inhibition test on Day 5. Thereafter, the prolonged BDNF mRNA and protein expression levels were measured in the hippocampus and the prefrontal cortex. Results – CIP dose-dependently reduced both locomotion and reward-motivated exploratory activity, accompanied with impaired learning ability. In contrast, there were no significant differences in startle reflex and sensory gating among treatment groups; however, CIP treatment reduced motor activity of the animals in this test, too. These alterations were associated with reduced BDNF mRNA and protein expression levels in the hippocampus but not the prefrontal cortex. Conclusion – This study revealed the detrimental effects of CIP treatment on locomotor activity and motivation/learning ability during osteoarthritic condition, which might be due to, at least partially, deficient hippocampal BDNF expression and ensuing impairments in neural and synaptic plasticity

Automating, analyzing and improving pupillometry with machine learning algorithms

The investigation of the pupillary light reflex (PLR) is a well-known method to provide information about the functionality of the autonomic nervous system. Pupillometry, a non-invasive technique, was applied to study the PLR alterations in a new, schizophrenia-like rat substrain, named WISKET. The pupil responses to light impulses were recorded with an infrared camera; the videos were automatically processed and features were extracted from the pupillograms. Besides the classical statistical analysis (ANOVA), feature selection and classification were applied to reveal the significant differences in the PLR parameters between the control and WISKET animals. Based on these results, the disadvantages of this method were analyzed and the measurement setup was redesigned and improved. The pupil segmentation method has also been adapted to the new videos. 2564 images were annotated manually and used to train a fully-convolutional neural network to produce pupil mask images. The method was evaluated on 329 test images and achieved 4% median relative error. With the new setup, the pupil detection became reliable and the new data acquisition offers robustness to the experiments

Feature extraction and classification for pupillary images of rats

The investigation of the pupillary light reflex (PLR) is a well-known method to provide information about the functionality of the autonomic nervous system. Pupillometry, a non-invasive technique, was applied in our lab to study the schizophrenia-related PLR alterations in a new selectively bred rat substrain, named WISKET. The pupil responses to light impulses were recorded with an infrared camera; the videos were automatically processed and features were extracted. Besides the classical statistical analysis (ANOVA), feature selection and classification were applied to reveal the significant differences in the PLR parameters between the control and WISKET animals