OBJECTIVE: To investigate the effects of CACNA1H gene knockout (KO) on autistic-like behaviors and the morphology of hippocampal neurons in mice. METHODS: In the study, 25 CACNA1H KO mice of 3-4 weeks old and C57BL/6 background were recruited as the experimental group, and 26 wild type (WT) mice of the same age and background were recruited as the control group. Three-chamber test and open field test were used to observe the social interaction, anxiety, and repetitive behaviors in mice. After that, their brain weight and size were measured, and the number of hippocampal neurons were observed by Nissl staining. Furthermore, the CACNA1H heterozygote mice were interbred with Thy1-GFP-O mice to generate CACNA1H^-/--Thy1⁺(KO-GFP) and CACNA1H^+/+-Thy1⁺ (WT-GFP) mice. The density and maturity of dendritic spines of hippocampal neurons were observed. RESULTS: In the sociability test session of the three-chamber test, the KO mice spent more time in the chamber of the stranger mice than in the object one (F_{1, 14}=95.086, P < 0.05; Post-Hoc: P < 0.05), without any significant difference for the explored preference index between the two groups (t=1.044, P>0.05). However, in the social novelty recognition test session, no difference was observed between the time of the KO mice spend in the chamber of new stranger mice and the stranger one (F_{1, 14}=18.062, P < 0.05; Post-Hoc: P>0.05), and the explored preference index of the KO mice was less than that of the control group (t=2.390, P < 0.05). In the open field test, the KO mice spent less time in the center of the open field apparatus than the control group (t=2.503, P < 0.05), but the self-grooming time was significantly increased compared with the control group (t=-2.299, P < 0.05). Morphological results showed that the brain weight/body weight ratio (t=0.356, P>0.05) and brain size (t=-0.660, P>0.05) of the KO mice were not significantly different from those of the control group, but the number of neurons were significantly reduced in hippocampal dentate gyrus compared with the control group (t=2.323, P < 0.05). Moreover, the density of dendritic spine of dentate gyrus neurons in the KO-GFP mice was significantly increased compared with the control group (t=-2.374, P < 0.05), without any significant difference in spine maturity (t=-1.935, P>0.05). CONCLUSION CACNA1H KO mice represent autistic-like behavior, which may be related to the decrease in the number of neurons and the increase in the density of dendritic spine in the dentate gyrus.
Animals ; Autistic Disorder/genetics* ; Calcium Channels, T-Type/genetics* ; Gene Knockout Techniques ; Hippocampus ; Mice ; Mice, Inbred C57BL ; Mice, Knockout ; Neurons

The thalamus is a brain structure known to modulate sensory information before relaying to the cortex. The unique ability of a thalamocortical (TC) neuron to switch between the high frequency burst firing and single spike tonic firing has been implicated to have a key role in sensory modulation including pain. Of the two firing modes, burst firing, especially maintaining certain burst firing properties, was suggested to be critical in controlling nociceptive behaviors. Therefore, understanding the factors that influence burst firing properties would offer important insight into understanding sensory modulation. Using computational modeling, we investigated how the balance of excitatory and inhibitory inputs into a TC neuron influence TC bursting properties. We found that intensity of inhibitory inputs and the timing of excitatory input delivery control the dynamics of bursting properties. Then, to reflect a more realistic model, excitatory inputs delivered at different dendritic locations—proximal, intermediate, or distal—of a TC neuron were also investigated. Interestingly, excitatory input delivered into a distal dendrite, despite the furthest distance, had the strongest influence in shaping burst firing properties, suggesting that not all inputs equally contribute to modulating TC bursting properties. Overall, the results provide computational insights in understanding the detailed mechanism of the factors influencing temporal pattern of thalamic bursts.
Brain ; Calcium Channels, T-Type ; Computational Biology ; Dendrites ; Fires ; Neurons ; Sensory Gating ; Thalamus

T-type calcium channels are low voltage-activated calcium channels that evoke small and transient calcium currents. Recently, T-type calcium channels have been implicated in neurodevelopmental disorders such as autism spectrum disorder and neural tube defects. However, their function during embryonic development is largely unknown. Here, we investigated the function and expression of T-type calcium channels in embryonic neural progenitor cells (NPCs). First, we compared the expression of T-type calcium channel subtypes (CaV3.1, 3.2, and 3.3) in NPCs and differentiated neural cells (neurons and astrocytes). We detected all subtypes in neurons but not in astrocytes. In NPCs, CaV3.1 was the dominant subtype, whereas CaV3.2 was weakly expressed, and CaV3.3 was not detected. Next, we determined CaV3.1 expression levels in the cortex during early brain development. Expression levels of CaV3.1 in the embryonic period were transiently decreased during the perinatal period and increased at postnatal day 11. We then pharmacologically blocked T-type calcium channels to determine the effects in neuronal cells. The blockade of T-type calcium channels reduced cell viability, and induced apoptotic cell death in NPCs but not in differentiated astrocytes. Furthermore, blocking T-type calcium channels rapidly reduced AKT-phosphorylation (Ser473) and GSK3β-phosphorylation (Ser9). Our results suggest that T-type calcium channels play essential roles in maintaining NPC viability, and T-type calcium channel blockers are toxic to embryonic neural cells, and may potentially be responsible for neurodevelopmental disorders.
Apoptosis ; Astrocytes ; Autism Spectrum Disorder ; Brain ; Calcium ; Calcium Channels ; Calcium Channels, T-Type* ; Cell Death ; Cell Survival ; Embryonic Development ; Female ; Neural Tube Defects ; Neurodevelopmental Disorders ; Neurons ; Pregnancy ; Stem Cells*

OBJECTIVETo investigate the rebound depolarization of substantia gelatinosa (SG) neurons in rat spinal dorsal horn and explore its modulatory mechanisms to provide better insights into rebound depolarization-related diseases.

METHODSParasagittal slices of the spinal cord were prepared from 3- to 5-week-old Sprague-Dawley rats. The electrophysiologic characteristics and responses to hyperpolarization stimulation were recorded using whole-cell patch-clamp technique. The effects of hyperpolarization-activated cyclic nucleotide gated cation (HCN) channel blockers and T-type calcium channel blockers on rebound depolarization of the neurons were studied.

RESULTSA total of 63 SG neurons were recorded. Among them, 23 neurons showed no rebound depolarization, 19 neurons showed rebound depolarization without spikes, and 21 neurons showed rebound depolarization with spikes. The action potential thresholds of the neurons without rebound depolarization were significantly higher than those of the neurons with rebound depolarization and spikes (-28.7∓1.6 mV vs -36.0∓2.0 mV, P<0.05). The two HCN channel blockers CsCl and ZD7288 significantly delayed the latency of rebound depolarization with spike from 45.9∓11.6 ms to 121.6∓51.3 ms (P<0.05) and from 36.2∓10.3 ms to 73.6∓13.6 ms (P<0.05), respectively. ZD7288 also significantly prolonged the latency of rebound depolarization without spike from 71.9∓35.1 ms to 267.0∓68.8 ms (P<0.05). The T-type calcium channel blockers NiCl2 and mibefradil strongly decreased the amplitude of rebound depolarization with spike from 19.9∓6.3 mV to 9.5∓4.5 mV (P<0.05) and from 26.1∓9.4 mV to 15.5∓5.0 mV (P<0.05), respectively. Mibefradil also significantly decreased the amplitude of rebound depolarization without spike from 14.3∓3.0 mV to 7.9∓2.0 mV (P<0.05).

CONCLUSIONNearly two-thirds of the SG neurons have rebound depolarizations modulated by HCN channel and T-type calcium channel.

Action Potentials ; Animals ; Calcium Channel Blockers ; pharmacology ; Calcium Channels, T-Type ; Cell Polarity ; Cesium ; pharmacology ; Chlorides ; pharmacology ; Cyclic Nucleotide-Gated Cation Channels ; antagonists & inhibitors ; Neurons ; cytology ; Patch-Clamp Techniques ; Pyrimidines ; pharmacology ; Rats ; Rats, Sprague-Dawley ; Spinal Cord Dorsal Horn ; cytology ; Substantia Gelatinosa ; cytology

OBJECTIVETo investigate the effect of KN93, a calmodulin-dependent protein kinase II (CaMK II) inhibitor, on SH-SY5Y cell injury induced by bupivacaine hydrochloride.

METHODSSH-SY5Y cells exposed for 24 h to 1 mmol/L KN93, 1 mmol/L bupivacaine hydrochloride, or both were examined for morphological changes and Cav3.1 protein expressions using Western blotting. The vitality and apoptosis rate of the cells at different time points during the exposures were assessed with MTT assay and flow cytometry, respectively.

RESULTSBupivacaine hydrochloride exposure caused obvious cell morphologial changes, reduced cell viability, increased cell apoptosis, and enhanced Cav3.1 protein expression. All these changes were partly reversed by treatment of the cells with 1 mmol/L KN93.

CONCLUSIONSCaMKII may play a role in bupivacaine hydrochloride-induced SH-SY5Y cells injury, which is related with upregulated Cav3.1 protein expression.

Apoptosis ; Bupivacaine ; adverse effects ; Calcium Channels, T-Type ; metabolism ; Calcium-Calmodulin-Dependent Protein Kinase Type 2 ; antagonists & inhibitors ; metabolism ; Cell Line ; Cell Survival ; Humans ; Up-Regulation

OBJECTIVE: To explore the expression levels of three T-type calcium channel receptors (α1G; α1H; α1I) in the cochlea and spiral ganglion neurons of C57BL/6J mice with different ages. METHOD: Thirty cases of C57BL/6J mice were divided into three groups (6-8 W, 24-26 W, 42-44 W) according to the age. The expressions of three T-type calcium channel receptors were quantified by RT-PCR after hearing thresholds measured by ABR. RESULT: Three receptors were detected in the cochlea and spiral ganglion neurons of 6-8 W C57BL/6J mice. The quantitative results showed that the expression levels of α1H and α1I were highest among three receptors in spiral ganglion neurons and in the cochlea respectively. The expression levels of three receptors significantly decreased with age,especially at the age of 4244 W. CONCLUSION The expression of T-type calcium channel receptors reduced with age in the inner ear of C57BL/6J mice.
Animals ; Calcium Channels ; Calcium Channels, T-Type ; biosynthesis ; Cochlea ; metabolism ; Ear, Inner ; Mice ; Mice, Inbred C57BL ; Neurons ; Receptors, Calcium-Sensing ; Spiral Ganglion ; metabolism

BACKGROUND AND OBJECTIVES: The aim of this study was to evaluate the effects of an L- and T-type calcium channel blocker (CCB) on 24-hour systolic blood pressure (24-hour SBP) and heart rate (24-hour HR) profiles in essential hypertensive patients. SUBJECTS AND METHODS: Thirty-seven consecutive patients were enrolled in this study. The 24-hour SBP and HR were recorded before and after treatment with efonidipine (L- and T-type CCB, 40 mg), after waking. Changes in 24-hour SBP and HR and the diurnal to nocturnal SBP ratio were measured. The best-fit curves of changes in SBP and HR were depicted using a periodic function. RESULTS: The mean 24-hour SBP and HR decreased significantly after treatment. The diurnal to nocturnal SBP ratio in dipper-type hypertension cases decreased from 16.7+/-6.1% to 8.3+/-9.8% (p<0.05), whereas in non-dipper hypertension cases, it increased from 2.3+/-2.9% to 7.7+/-5.1% (p<0.01). The antihypertensive effect was minimal at 5.0 hours after drug administration and it slowly recovered at a constant rate (2.1 mm Hg/h) over 12 hours in dipper cases. The median 24-hour changes in HR in the dipper and non-dipper cases were -2.3/min and -5.4/min, respectively. A continuous reduction in the change in HR was seen from 3.5 to 23 hours after drug administration. CONCLUSION: The antihypertensive action of efonidipine was characterized by a slow recovery of the SBP decrease at a constant rate (2.1 mm Hg/h) and a non-administration time dependent reduction in 24-hour HR.
Blood Pressure ; Calcium Channel Blockers ; Calcium Channels, T-Type ; Dihydropyridines ; Heart ; Heart Rate ; Humans ; Hypertension ; Nitrophenols ; Organophosphorus Compounds

OBJECTIVETo observe the anti-remodeling effect of acupuncture on asthma and to explore the mechanism of T-type calcium channel protein in airway smooth muscle cell in airway remodeling effect in asthma.

METHODSThirty-two rats were randomly divided into a normal group, a model group, an acupuncture group and a sham acupuncture group, 8 rats in each group. The rats in the latter three groups were sensitized for consecutive 14 days by single peritoneal injection of aqueous suspension 1 mL of 10 mg ovalbumin (OVA), 200 mg aluminum hydroxide and saline together with 1 mL inactivated pertussis vaccine. From the 15th day, asthma was induced for 30 minutes by ultrasonic atomizing inhalation of 1% OVA for consecutive 14 days in the model group. The acupuncture group was treated with acupuncture at "Dazhui" (GV 14), "Fengmen" (BL 12) and "Feishu" (BL 13) for 30 minutes before the ultrasonic atomizing inhalation, once every two days for consecutive 14 days. The same acupoints selection and the course of treatment as the acupuncture group were produced in the sham acupuncture group and they were treated with acupuncture at 1 mm acupoint skin without retaining needles. The normal group remained unhandled. The respiratory function and the airway remodeling were evaluated by airway resistance and pulmonary histopathology, respectively, and the T-type calcium channel protein expression of Ca(v)3.1, Ca(v) 3.2, Ca, 3.3 in airway smooth muscle cell were detected by immunohistochemistry technique.

RESULTS(1) The airway resistance in the model group was higher than that in the normal group and in the acupuncture group (both P < 0.05), and the airway resistance in the acupuncture group was lower than that in the sham acupuncture group (P < 0.05). (2) The ratios of the airway wall thickness to the basement membrane perimeter (Awt/Pbm) and the airway outer perimenter to the airway internal perimeter (Po/Pi) in the model group were higher than those in the normal group and in the acupuncture group (all P < 0.05), and the ratios of Awt/Pbm and Po/Pi in the acupuncture group were lower than those in the sham acupuncture group (both P < 0.05). (3) The average optical of Ca(v) 3.1 and Ca(v) 3.2 in airway smooth muscle cell in the model group were higher than that in the normal group and in the acupuncture group (both P < 0.05), and the average optical of Ca(v) 3.3 in airway smooth muscle cell in the model group was higher than that in the normal group (P < 0.05) and it was lower than that in the sham acupuncture group (P < 0.05).

CONCLUSIONAcupuncture can inhibit the airway remodeling and the accrementition of the airway smooth muscle and can reduce the airway resistance. The mechanism may be related to the inhibition of T-type calcium channel protein in airway smooth muscle cell, especially in relation to the protein expression of Ca(v) 3.1.

Acupuncture Therapy ; Airway Remodeling ; Animals ; Asthma ; genetics ; metabolism ; physiopathology ; Calcium Channels, T-Type ; genetics ; metabolism ; Humans ; Male ; Myocytes, Smooth Muscle ; metabolism ; Rats ; Rats, Sprague-Dawley ; Respiratory System ; metabolism ; physiopathology

OBJECTIVETo study the relationship of varicocele (VC) with the expressions of T-type channel alpha1H and alpha1G in the sperm of VC patients.

METHODSBased on the WHO criteria, we examined the semen samples by computer-aided sperm analysis (CASA), and divided the samples into groups A (normal semen from volunteers, n = 20), B (normal semen from VC patients, n = 16) and C (abnormal semen from VC patients, n = 44). We optimized the semen by discontinuous Percoll grade centrifugation, and determined the mRNA expressions of T-type channel alpha1H and alpha1G in the three groups using using reverse transcription polymerase chain reaction (RT-PCR).

RESULTSCompared with group A, the mRNA expressions of alpha1H and alpha1G showed with no significant decrease in group B (P>0.05), but were remarkably reduced in group C (P<0.01).

CONCLUSIONThe abnormal mRNA expressions of T-type channel alpha1H and alpha1G may be one of the causes of declined semen quality and consequently infertility in VC patients, which has pointed out a new direction for the studies of the causes and treatment of VC-related infertility.

Adolescent ; Adult ; Calcium Channels, T-Type ; genetics ; metabolism ; Case-Control Studies ; Humans ; Infertility, Male ; genetics ; Male ; RNA, Messenger ; genetics ; Semen Analysis ; Spermatozoa ; metabolism ; Varicocele ; genetics ; metabolism ; Young Adult

This article reports the investigation of the effect of carvedilol (Car) on T-type calcium current (I(Ca,T)) of noninfarcted ventricular myocytes in rabbit models of healed myocardial infarction (HMI). Rabbits with left anterior descending artery ligation were prepared and allowed to recover for 8 weeks, as HMI group. Animals undergoing an identical surgical procedure without coronary ligation were served as the sham-operated group (sham group). Whole cell voltage-clamp techniques were used to measure and compare currents in cells from the different groups. Noting that I(Ca,T) density in HMI cells increased markedly to -2.36 +/- 0.12 pA/pF (at -30 mV) compared with cells of sham, where little I(Ca,T) (-0.35 +/- 0.02 pA/pF) was observed. Meanwhile, further analysis revealed a significant hyperpolarizing shift of steady-state activation curve of I(Ca,T) in HMI cells, where the time constants of deactivation were prolonged and the time of recovery from inactivation was shortened. Finally, the amplitude of I(Ca,T) was increased. Carvedilol (1 micromol x L(-1)) was found to decrease the amplitude of I(Ca,T) to -1.38 +/- 0.07 pA/pF through inhibiting process of I(Ca,T) activation. Furthermore, carvedilol delayed recovery from inactivation of I(Ca,T) and shortened the time constants of deactivation in HMI cells. This study suggested that the application of carvedilol in HMI cells contributes to the dynamic changes in I(Ca,T) and may account for reduction of incidence of arrhythmia after myocardial infarction.
Adrenergic beta-Antagonists ; pharmacology ; Animals ; Calcium Channels, T-Type ; drug effects ; Carbazoles ; pharmacology ; Female ; Male ; Myocardial Infarction ; pathology ; physiopathology ; Myocytes, Cardiac ; drug effects ; physiology ; Patch-Clamp Techniques ; Propanolamines ; pharmacology ; Rabbits