| Schizophrenia is a kind of clinical neurodegenerative diseases with higher incidence and lower cure rate, its pathogenesis is associated with many substances which become abnormal, such as dopamine (DA),5-serotonin and glutamate. In which, the influence of the imbalance of DA neurotransmitter release on schizophrenia is a theory which is common recognized. At present, It has been reported by the research on neuroanatomy that midbrain DA neurons project to the different areas of the brain during development, forming different subsets of neurons which have different functions. In which, mesocortical DA neurons located in the ventral tegmental area of the midbrain project to the prefrontal cortex of the brain, which forms the mesocortical DA pathway. The abnormal of the pathway is the anatomical basis for leading to schizophrenia and other psychiatric disorders. Researches show that the mesocortical DA neurons with electrophysiological specific characteristics. In most cases spontaneous discharge frequency is higher than the mesostriatal DA neurons, and in the basis of increasing injection amplitude of the hyperpolarizing current, mesocortical DA neurons show no obvious "sag". Mesencephalic DA neurons can release DA from somata or dendrites of its own in the substantia nigra and VTA, activating D2autoreceptor to inhibit the firing of the cell and reduce the release of DA. However, the regulation of electrophysiological characteristics and DA release of the mesocortical DA neurons which project to the prefrontal cortex is still not clear so far.Aripiprazole, as a new type of antipsychotic drugs, can effectively alleviate the symptoms of schizophrenia patients, and have neuroprotective properties. Clinical studies show that, in addition to activating5-HT1A receptor, antagonistic with5-HT2A, Aripiprazole can also partially agitate on DA D2receptor. Recently reported in the literature, aripiprazole could inhibit DA release induced by the high K+solution in PC12cells. Recording of the cells in vivo shows that aripiprazole can reduce the activity of the spontaneous firing of the VTA in vivo. Activation of G protein-coupled inwardly rectifying potassium (GIRK) can inhibit DA release of the neurons in the VTA and substantia nigra. At present, there is no specific reports on whether aripiprazole play a role in the treatment of schizophrenia through affect on DA secretion and the electrophysiological properties of mesocortical DA neurons.Based on the idea of the studies above, we speculated that the mechanism of aripiprazole in the treatment of schizophrenia, which might have some relationship with the affection of the mesocortical DA neurons electrophysiological characteristics and DA release. In order to confirm this idea, this experiment was conducted the following experiments:1. According to stereotaxic instrument, we used neuronal retrograde tracing technique to put fluorescent tracers accurately injected to the prefrontal cortex in the brain of the rats, which was the projecting target of the mesocortical DA neurons. Through axoplasmic retrograde transport, we could label the mesocortical DA neurons which projected to the target area. And perform tyrosine hydroxylase (TH) immunohistochemistry staining to determine;2. Taking up the midbrain tissue labeled to make the brain slices, we recorded the amplitude, frequency, width, and detected the changes of the resting membrane potential and threshold through the glass electrodes with different concentrations of aripiprazole on spontaneous and evoked action potentials of the mesocortical DA neurons using brain slice patch-clamp technique in the whole-cell recording mode.3. Detect the effect of aripiprazole on the changes of the outward K+current of the mesocotical DA neurons, then analysis the possible mechanism of aripiprazole in modulating the excitability of the mesocortical DA neurons.4. In the tissue slices prepared of the midbrain, we detected the affection of aripiprazole on DA release of the mesocortical DA neurons using carbon fiber electrode technique. In the stimulation of the780mV voltage, carbon fiber could oxidate DA to dopamine quinone. DA oxidation current could be detected by the carbon fiber electrode and transformed the changes of the DA qualitative into voltammogram, which could achieve to the nanomolar and sub-second accuracy; and applying with the DA standard liquid to detect the situation of carbon fiber electrode.5. In vivo, after the injection of aripiprazole in the VTA of the brain, using carbon fiber electrode technology to detect the changes of DA release in the prefrontal cortex which the mesocortical DA neurons projected to.Results showed:1. Fluorescent tracer could accurately lable the mesocortical DA neurons which projected to the prefrontal cortex. Green fluorescent tracer distributed in cytoplasm of neurons, and it also could be found in the axon of the neurons as the tracer particles. Immunohistochemical staining for TH showed that labeled cells were TH-positive, and mainly located in the ventral tegmental area.2. We detected the affect of aripiprazole in the different concentrations on the spontaneous action potentials and evoked action potentials of the mesocortical DA neurons labeled using brain slices patch-clamp technique. Add10μM aripiprazole to the perfusate, the frequency (P<0.05) and amplitude (P<0.01) of spontaneous action potentials was significantly reduced. Compared with dosing ago, resting potential and threshold values were significantly lower (P<0.05); the current stimulus of (300pA,500ms) were given to labeled cells, and the action potentials were evoked at frequency of30±1.15Hz, and amplitude of55.5±0.64mV.10μM aripiprazole significantly reduced their frequency and amplitude (P<0.05). High concentrations of aripiprazole (100μM) made the spontaneous and evoked action potentials disappeared. These results showed that aripiprazole could inhibit the excitability of the mesocortical DA neurons.3. We performed the detection on external K+current to further analysis the mechanism of aripiprazole in suppressing excitability of mesocortical DA neurons. Add10μM aripiprazole to the perfusate, potassium current could increase through stimulation of10μM aripiprazole on the mesocortical DA neurons (P<0.05); with the perfusate washed away, the potassium current recovered.4. With the addition of aripiprazole in the perfusate of the brain slices, the oxidation current produced by the DA release of labeled DA neurons could be detected by the carbon fiber electrode. Compared with that before dosing, amplitude (P<0.05) and the area under the curve (P<0.01) of the current were significantly decreased, indicating that aripiprazole could inhibit DA release of the mesocortical DA neurons labeled in the brain slices. Using70mM KC1as positive control, when the membrane holding potential of the electrode transited to780mV, there would be a formed peak on the oxidation current spectrum detected by the carbon fiber electrode; When the holding potential returned to0mV, the peak disappeared.5. In vivo, with the injection of10μM aripiprazole to the VTA, the oxidation current of the DA release of the prefrontal cortex which the mesocortical DA neurons projected to could be detected by the carbon fiber electrode. After adding aripiprazole for2h, the oxidation current of DA in the prefrontal cortex reduced; and it would become to the lowest when adding aripiprazole for3h. Until8h, the current would be back to the level before dosing.In summary, the experimental results show that aripiprazole can inhibit spontaneous and evoked action potentials of the mesocortical DA neurons through excitation of the outward K+current. Further, it can inhibit the excitability of the mesocortical DA neurons, and reduce the secretion of DA of the mesocortical DA neurons to participate in the treatment of schizophrenia. |
PDF Full Text Request