Action Sequence Learning Is Impaired in Genetically Modified Mice with the Suppressed GABAergic Transmission from the Thalamic Reticular Nucleus to the Thalamus.

The thalamic reticular nucleus (TRN) is a thin sheet of GABAergic neurons surrounding the thalamus, and it regulates the activity of thalamic relay neurons. The TRN has been reported to be involved in sensory gating, attentional regulation, and some other functions. However, little is known about the contribution of the TRN to sequence learning. In the present study, we examined whether the TRN is involved in reward-based learning of action sequence with no eliciting stimuli (operant conditioning), by analyzing the performance of male and female Avp-Vgat-/- mice (Vgatflox/flox mice crossed to an Avp-Cre driver line) on tasks conducted in an operant box having three levers. Our histological and electrophysiological data demonstrated that in adult Avp-Vgat-/- mice, vesicular GABA transporter (VGAT) was absent in most TRN neurons and the GABAergic transmission from the TRN to the thalamus was largely suppressed. The performance on a task in which mice needed to press an active lever for food reward showed that simple operant learning of lever pressing and learning of win-stay and lose-shift strategies are not affected in Avp-Vgat-/- mice. In contrast, the performance on a task in which mice needed to press three levers in a correct order for food reward showed that learning of the order of lever pressing (action sequence learning) was impaired in Avp-Vgat-/- mice. These results suggest that the TRN plays an important role in action sequence learning.

Evaluation of motor learning in predictable loading task using a force sense presentation device.

Neural circuits connecting the cerebellum with the cerebral cortex are important for both motor and cognitive functions. Therefore, assessment of cerebellar function is clinically important for patients with various motor and cognitive dysfunctions. Cerebellum-dependent motor learning has been studied using various tasks. The most widely used tasks are visuomotor adaptation tasks, in which subjects are required to make movements in two dimensions. Studies using simpler tasks of one-dimensional movement, which are easier for patients with motor problems to perform, have suggested that anticipatory responses in these tasks are useful to evaluate cerebellum-dependent motor control or motor learning. In this study, we examined whether the motor learning process can be evaluated in a simple loading task. Using space interface device for artificial reality (SPIDAR), a constant downward force was loaded to subjects' hands in a predictable condition, and the vertical movement of the hand was recorded. The hand deflection from the initial position was displayed on a screen for visual feedback information. We examined effects of repeated loading task training (90 times) on hand movements, by analyzing a small upward movement just before loading (anticipatory response) and a large downward movement after loading in each trial. We found that the repeated training lowered the time constant of upward movement and reduced the amplitude and time-to-peak of downward movement. These training effects were maintained into the next day. Furthermore, we found that loading task training with eyes closed was also effective, which indicates that proprioceptive information is enough for improvement of performance.

Effects of environmental enrichment on exploratory behavior, win-stay and lose-shift performance, motor sequence learning, and reversal learning during the three-lever operant task in mice.

Beneficial effects of environmental enrichment (EE) on the central nervous system have been demonstrated. Although the effects of EE on spatial learning have been extensively studied, studies on reward-based motor learning are limited. In this study we examined the effects of EE on the performance of operant tasks using three levers (A-C). Mice were divided into two groups and housed either in the control condition or in the physical EE condition. The mice were trained in three types of operant tasks in sequence. First, mice were trained to press one of the active levers for a food reward (one-lever task). Second, mice were trained to press the three levers in the order of A, B, and C (three-lever task). Third, the lever order was reversed to C, B, and A (reverse three-lever task). We found some behavioral differences between control and EE mice. When all three levers were active in the one-lever task, mice tended to press the three levers equally at first, then shifted to press one lever preferentially. This behavioral shift from exploration to exploitation was delayed in EE mice. When only one lever was active, EE mice showed a higher lose-shift performance. In the three-lever and reverse three-lever tasks, EE mice pressed three levers more often and acquired more food rewards, compared to control mice, although the success rate in both tasks was not different between the two groups. These behavioral features observed in EE mice (higher lose-shift performance and higher trial and error activity) might be advantageous when circumstances are not stable.

The use of a cold pack during resistance exercises is effective for reducing intramuscular oxygenation and increasing myoelectric activity.

[Purpose] The purpose of this study was to determine the efficacy of using a cold pack while doing resistance exercises for enhancing muscle strength and muscle hypertrophy through decreased intramuscular oxygenation and/or increased myoelectric activity. [Participants and Methods] Twenty-four resistance-trained males (age: 26.4 ± 8.4 years, height: 169.3 ± 5.2 cm, body weight: 74.7 ± 8.8 kg) involved in this study. All the participants completed two experimental sessions in random order (cold pack resistance exercise and resistance exercise) with a 3-day interval. Four types of resistance exercises (4 sets × 8 repetitions with an 8-repetition maximum) targeting the right triceps brachii muscle were performed in both the experimental sessions. [Results] The percentage baseline oxyhemoglobin/myoglobin level during resistance exercise was significantly lower, the half-recovery time of muscle oxygenation in intervals between sets was significantly longer, and the myoelectric activity was significantly higher in the cold pack resistance exercise than in the resistance exercise session. [Conclusion] The results suggest that using a cold pack with resistance exercises is effective in inducing intramuscular deoxygenation and increasing myoelectric activity and may be useful for increasing muscle strength and inducing hypertrophy.

Endocannabinoid Signaling from 2-Arachidonoylglycerol to CB1 Cannabinoid Receptor Facilitates Reward-based Learning of Motor Sequence.

The endocannabinoid system modulates synaptic transmission, controls neuronal excitability, and is involved in various brain functions including learning and memory. 2-arachidonoylglycerol, a major endocannabinoid produced by diacylglycerol lipase-α (DGLα), is released from postsynaptic neurons, retrogradely activates presynaptic CB1 cannabinoid receptors, and induces short-term or long-term synaptic plasticity. To examine whether and how the endocannabinoid system contributes to reward-based learning of a motor sequence, we subjected male CB1-knockout (KO) and DGLα-KO mice to three types of operant lever-press tasks. First, we trained mice to press one of three levers labeled A, B, and C for a food reward (one-lever task). Second, we trained mice to press the three levers in the order of A, B, and C (three-lever task). Third, the order of the levers was reversed to C, B, and A (reverse three-lever task). We found that CB1-KO mice and DGLα-KO mice exhibited essentially the same deficits in the operant lever-press tasks. In the one-lever task, both strains of knockout mice showed a slower rate of learning to press a lever for food. In the three-lever task, both strains of knockout mice showed a slower rate of learning of the motor sequence. In the reverse three-lever task, both strains of knockout mice needed more lever presses for reversal learning. These results suggest that the endocannabinoid system facilitates reward-based learning of a motor sequence by conferring the flexibility with which animals can switch between strategies.

NPC1L1-dependent intestinal cholesterol absorption requires ganglioside GM3 in membrane microdomains.

Intestinal cholesterol absorption is a key regulator of systemic cholesterol homeostasis. Excessive dietary cholesterol and its intestinal uptake lead to hypercholesterolemia, a major risk factor for cardiovascular disease. Intestinal cholesterol uptake is mediated by Niemann-Pick C1-like 1 (NPC1L1), a transmembrane protein localized in membrane microdomains (lipid rafts) enriched in gangliosides and cholesterol. The roles of gangliosides, such as monosialodihexosylganglioside (GM3) and its synthesizing enzyme GM3 synthase (GM3S), in NPC1L1-dependent cholesterol uptake have not been examined previously. Here, we examined NPC1L1-dependent cholesterol uptake in a cell model as well as in wild-type and apoE-deficient mice fed normal or high-cholesterol diets. We showed that NPC1L1-dependent cholesterol uptake was impaired in GM3S-deficient cells and that GM3S deficiency promoted resistance to hypercholesterolemia in both wild-type and apoE-deficient mice fed the high-cholesterol but not the normal diet. Our findings suggest that GM3 and related gangliosides are essential for NPC1L1-mediated intestinal cholesterol absorption and are potential targets for hypercholesterolemia therapy.

Electrophysiological evidence showing muscarinic agonist-antagonist activities of N-desmethylclozapine using hippocampal excitatory and inhibitory neurons.

The atypical antipsychotic clozapine is widely used for treatment-resistant schizophrenic patients. Clozapine and its major active metabolite, N-desmethylclozapine (NDMC), have complex pharmacological properties, and interact with various neurotransmitter receptors. There are several biochemical studies reporting that NDMC exhibits a partial agonist profile at the human recombinant M1 muscarinic receptors. However, direct electrophysiological evidence showing the ability of NDMC to activate native M1 receptors in intact neurons is poor. Using rat hippocampal neurons, we previously demonstrated that activation of muscarinic receptors by a muscarinic agonist, oxotremorine M (oxo-M), induces a decrease in outward K(+)current at -40mV. In the present study, using this muscarinic current response we assessed agonist and antagonist activities of clozapine and NDMC at native muscarinic receptors in intact hippocampal excitatory and inhibitory neurons. Suppression of the oxo-M-induced current response by the M1 antagonist pirenzepine was evident only in excitatory neurons, while the M3 antagonist darifenacin was effective in both types of neurons. Muscarinic agonist activity of NDMC was higher than that of clozapine, higher in excitatory neurons than in inhibitory neurons, sensitive to pirenzepine, and partially masked when co-applied with clozapine. Muscarinic antagonist activity of clozapine as well as NDMC was not different between excitatory and inhibitory neurons, but clozapine was more effective than NDMC. These results demonstrate that NDMC has the ability to activate native M1 receptors expressed in hippocampal excitatory neurons, but its agonist activity might be limited in clozapine-treated patients because of the presence of excessive clozapine with muscarinic antagonist activity.

[Analysis of brain activity during delayed matching-to-sample test].

Despite recent advances in uncovering the neural signature of memory processing in humans, spatiotemporal characteristics of cerebral activity during mental transformation of the visual (color) memory to the visuo-spatial memory has not been studied. The aim of this study was to investigate the electrophysiological correlates of internal memory transformation based on the event-related potential analysis using a root mean square (RMS) and a standardized low resolution electromagnetic tomography (sLORETA). Eleven healthy human subjects completed a modified delayed matching-to-sample test in which the visual(color) short-term memory had to be transformed into the visuo-spatial memory according to the matching probe. In comparison with a non-memory control condition, identifiable RMS peak was observed 352ms after the matching probe presentation in the condition of memory transformation. In this time, sLORETA demonstrated the higher current density in the prefrontal region. Our results suggested that the prefrontal region is associated with the internal memory transformation process depicting a spatiotemporal profile similar to P3b.

Effects of clozapine and N-desmethylclozapine on synaptic transmission at hippocampal inhibitory and excitatory synapses.

Clozapine is the first atypical antipsychotic, and improves positive and negative symptoms of many patients with schizophrenia resistant to treatment with other antipsychotic agents. Clozapine induces minimal extrapyramidal side effects, but is more often associated with seizures. A large number of studies have been conducted to elucidate pharmacological profiles of clozapine and its major active metabolite, N-desmethylclozapine (NDMC). However, there are only a limited number of electrophysiological studies examining their effects on synaptic transmission. In this study, we examined effects of clozapine and NDMC on synaptic transmission by measuring inhibitory and excitatory postsynaptic currents in rat cultured hippocampal neurons. We found that clozapine and NDMC have qualitatively similar actions. They depressed the inhibitory transmission at 1-30 µM, and the excitatory transmission at 30 µM, the former being much more sensitive. The depression of IPSCs by 30 µM of these drugs was associated with an increase in the paired-pulse ratio. The GABA-induced currents were suppressed by these drugs, but less sensitive than IPSCs. The AMPA-induced currents were slightly potentiated by these drugs at 30 µM. At 30 µM, clozapine and NDMC slightly suppressed Ca(2+) and Na(+) channels. These results strongly suggest that clozapine and NMDC depress the inhibitory synaptic transmission mainly by antagonizing postsynaptic GABA(A) receptors, but at higher concentrations additionally by acting on presynaptic site, possibly in part through inhibition of presynaptic Ca(2+) and Na(+) channels. Preferential depression of inhibitory synaptic transmission by clozapine and NDMC might contribute to therapeutic actions and/or side-effects of clozapine.