No evidence of a causal relationship between miscarriage and 25-hydroxyvitamin D: a Mendelian randomization study.

STUDY QUESTION: Is there a causal relationship between 25-hydroxyvitamin D (25OHD) and miscarriage? SUMMARY ANSWER: In this study, little evidence of a causal relationship was found between low serum 25OHD concentration or vitamin D deficiency and the risk of miscarriages. WHAT IS KNOWN ALREADY: Associations between low vitamin D levels and increased risk of miscarriage have been reported, but causality is unclear. STUDY DESIGN SIZE DURATION: The latest and largest genome-wide association studies (GWAS) for serum 25OHD concentration (n = 417 580), vitamin D deficiency (426 cases and 354 812 controls), miscarriage (16 906 cases and 149 622 controls), and the number of miscarriages (n = 78 700) were used to explore the causal association between serum vitamin D levels and miscarriage by two-sample Mendelian randomization analysis. PARTICIPANTS/MATERIALS SETTING METHODS: This study was based on summary GWAS results from the FinnGen database and the UK Biobank. The random-effect inverse-variance weighted method was regarded as the primary analysis; MR-Egger, weighted median, weighted mode, simple mode, and MR-pleiotropy residual sum and outlier (MR-PRESSO) were further employed as complementary methods. MR-Egger intercept analysis and MR-PRESSO were employed to test pleiotropy, and Cochran's Q statistic and leave-one-out sensitivity analysis were used to determine the heterogeneity and robustness of the overall estimates, respectively. MAIN RESULTS AND THE ROLE OF CHANCE: There was insufficient evidence of causal associations between serum 25OHD concentration and miscarriage (odds ratio (OR) = 0.995, 95% CI: 0.888 to 1.114, P = 0.927), or the number of miscarriages (ß = -0.004, 95% CI: -0.040 to 0.032, P = 0.829). Furthermore, little evidence of causality between genetically determined vitamin D deficiency to miscarriage (OR = 0.993, 95% CI: 0.966 to 1.021, P = 0.624), or the number of miscarriages (ß = 0.001, 95% CI: -0.009 to 0.011, P = 0.828), was observed. The results of the sensitivity analysis were robust, and no significant heterogeneity or horizontal pleiotropy was found. LIMITATIONS REASONS FOR CAUTION: This study is limited by the absence of female-specific GWAS data and the limited amount of GWAS data available for this study, as well as the need for caution in generalizing the findings to non-European ethnic groups. WIDER IMPLICATIONS OF THE FINDINGS: These findings enhance the current understanding of the intricate association between vitamin D and pregnancy outcomes, challenging prevailing beliefs regarding the strong association with miscarriage. The results provide a special perspective that may prompt further exploration and potentially offer insights for guiding future research and informing clinical guidelines pertaining to the management of miscarriage. STUDY FUNDING/COMPETING INTERESTS: This project was supported by the Hubei Provincial Natural Science Foundation Program General Surface Project (2022CFB200), the Key Research & Developmental Program of of Hubei Province (2022BCA042), the Fundamental Research Funds for the Central Universities (2042022gf0007, 2042022kf1210), and the Interdisciplinary Innovative Talents Foundation from Renmin Hospital of Wuhan University (JCRCWL-2022-001, JCRCYG-2022-009). All authors have no conflicts of interest to declare. TRIAL REGISTRATION NUMBER: N/A.

Effects of Tai Chi Chuan on Cognitive Function in Adults 60 Years or Older With Type 2 Diabetes and Mild Cognitive Impairment in China: A Randomized Clinical Trial.

Importance: Type 2 diabetes (T2D) is associated with the progression of cognitive dysfunction. Physical activity benefits cognition, but no evidence from randomized clinical trials has shown whether tai chi chuan has better long-term benefits than fitness walking in cognitive function for patients with T2D and mild cognitive impairment (MCI). Objective: To compare the effectiveness of tai chi chuan, a mind-body exercise, for improving cognitive function in older adults with T2D and MCI, with fitness walking. Design, Setting, and Participants: This randomized clinical trial was conducted between June 1, 2020, and February 28, 2022, at 4 sites in China. Participants included 328 adults (aged ≥60 years) with a clinical diagnosis of T2D and MCI. Interventions: Participants were randomized in a 1:1:1 ratio to a tai chi chuan group, a fitness walking group, or a control group. The tai chi chuan group received 24-form simplified tai chi chuan. The fitness walking group received fitness walking training. Both exercise groups took the training for 60 min/session, 3 times/wk, for 24 weeks in a supervised setting. All 3 groups were provided with a 30-minute diabetes self-management education session, once every 4 weeks for 24 weeks. The participants were followed up for 36 weeks. Main Outcomes and Measures: The primary outcome was the global cognitive function measured at 36 weeks by the Montreal Cognitive Assessment (MoCA). Secondary outcomes included MoCA at 24 weeks and other cognitive subdomain measures and blood metabolic indices at 24 and 36 weeks. Results: A total of 328 participants (mean [SD] age, 67.55 [5.02] years; mean [SD] T2D duration, 10.48 [6.81] years; 167 [50.9%] women) were randomized to the tai chi chuan group (n = 107), fitness walking group (n = 110), or control group (n = 111) and included in the intention-to-treat analysis. At 36 weeks, the tai chi chuan group showed improved MoCA scores compared with the fitness walking group (mean [SD], 24.67 [2.72] vs 23.84 [3.17]; between-group mean difference, 0.84 [95% CI, 0.02-1.66]; P = .046) in the intention-to-treat analysis. The per-protocol analysis data set and subgroup analysis at 36 weeks showed similar results. Based on the generalized linear models, the treatment effects were similar in each group after adjusting for self-reported dietary calories and physical activity. There were 37 nonserious adverse events (tai chi chuan group, 8; fitness walking group, 13; control group, 16) unrelated to the study with no statistically significant difference among the 3 groups (P = .26). Conclusions and Relevance: In this randomized clinical trial including older adults with T2D and MCI, tai chi chuan was more effective than fitness walking in improving global cognitive function. The findings support a long-term benefit, suggesting the potential clinical use of tai chi chuan as an exercise intervention to improve cognitive function for older adults with T2D and MCI. Trial Registration: ClinicalTrials.gov Identifier: NCT04416841.

Effects of Positive Psychological Nursing Combined with Free Posture on the Prognosis of Primipara with Singleton Spontaneous Delivery.

Objective: To explore the effects of positive psychological nursing combined with free posture on the prognosis of primipara with singleton spontaneous delivery. Methods: 106 cases of primipara with singleton spontaneous delivery who were admitted to the obstetrics department of our hospital from January 2017 to December 2019 were selected as the research subjects and they were divided into the observation group and control group according to the random number table method and maternal willingness, with 53 cases in each group. The control group received routine nursing, and the observation group was given positive psychological nursing combined with free posture on the basis of the control group. The stress degree during delivery (Hamilton anxiety scale (HAMA) and Hamilton depression scale (HAMD)), the time of first stage of labor, the time of second stage of labor, the time of third stage of labor, pain level during the active period of the first stage of labor, and delivery outcomes were observed and compared between the two groups. Results: The degree of anxiety and depression during childbirth in the observation group was significantly lower than that in the control group (P < 0.05). The time of first stage of labor and the time of second stage of labor in the observation group were shorter than those in the control group (P < 0.05), and there was no significant difference in the time of the third stage of labor (P > 0.05). The pain degree in the active stage of the first stage of labor in the observation group was lower than that in the control group (P < 0.05). The pain degree in the active stage of the first stage of labor in the observation group was lower than that in the control group (P < 0.05). Conclusion: Positive psychological nursing combined with free posture for primipara with singleton spontaneous delivery can shorten the stages of labor, reduce the pain, relieve psychological stress, and improve the delivery outcomes.

Mesenchymal stem cells moderate experimental autoimmune uveitis by dynamic regulating Th17 and Breg cells response.

Mesenchymal stem cells (MSCs) are adult stem cells from mesoderm with multi potential differentiation, and are being widely studied as a promising treatment for autoimmune diseases. The main inflammatory factors of experimental autoimmune uveitis (EAU) are T helper type 1 (Th1) and Th17. Regulatory B cells (Bregs) are a newly designated B cell subgroup, which has been proved to play a key role in regulating inflammation, autoimmunity and cancer. In this regard, we establish the EAU model by injecting interphotoreceptor retinoid-binding protein combined with complete Freund's adjuvant into the tail vein and bilateral thighs of rats, and inject MSCs or equal volume of phosphate buffer saline intraperitoneally on the day of immunization. Dynamic changes of cell subsets and cytokine expression are tested at different time periods to explore the relationship between MSCs treatment and disease prognosis during EAU course. Our results suggest that compared with the model control group, MSCs treatment can significantly reduce the production of Th1 and Th17 cytokines during EAU, while the production of regulatory B cells (Bregs) cytokines is significantly increased. At the same time, MSCs can reduce the proportion of Th17 in lymphocytes while the proportion of Bregs is elevated, thus inhibiting the differentiation and activity of interleukin in EAU rats. All this results provide more powerful evidence for cell therapy of autoimmune uveitis.

The association between periconceptional folic acid supplementation and the risk of preterm birth: a population-based retrospective cohort study of 200,000 women in China.

PURPOSE: The aim of this study was to examine the association between periconceptional folic acid (FA) supplementation and risk of preterm birth. METHODS: We conducted a retrospective cohort study in women aged 18-49 who participated in the NFPHEP from 2010 to 2018, and had a singleton livebirth in 129 counties in southwest China. Participants were divided into four groups according to the time period starting FA use: no use, after the last menstrual period, at least 1-2 months before the last menstrual period, at least 3 months before the last menstrual period. The outcomes were preterm birth (gestation < 37 weeks) and early preterm birth (gestation < 34 weeks). RESULTS: 201,477 women were included and 191,809 (95.2%) had taken FA during periconception. Compared with women who did not take FA, women who started taking FA 1-2 months before their last menstrual period had a 15% lower risk of preterm birth (aOR = 0.85, 95% CI 0.79-0.92), and women who started taking FA at least 3 months before their last menstrual period had a 20% lower risk of preterm birth (aOR = 0.80, 95% CI 0.75-0.87), but women who started taking FA after their last menstrual period did not appear to reduce the risk of preterm birth. CONCLUSIONS: In this study of 200,000 Chinese women, periconceptional supplementation with FA was associated with a lower risk of preterm birth. Women who started taking FA at least 3 months before their last menstrual period were more likely to reduce the risk of preterm birth.

Association between sleep duration in early pregnancy and risk of gestational diabetes mellitus: a prospective cohort study.

AIMS: As cohort studies of the impact of sleep duration during early pregnancy on gestational diabetes mellitus (GDM) are lacking, our study aimed to explore the association between sleep duration in the first trimester and GDM in one region of mainland China. METHODS: For this prospective cohort study, sleep duration data were collected from 3692 pregnant women at the first prenatal care appointment before 14 weeks of gestation. Multivariable log-binomial regression models were used to analyze the association of sleep duration with GDM after adjusting for demographic characteristics, health status (such as family history of diabetes, history of GDM, prepregnancy body mass index, gestational weight gain) and lifestyle habits (such as physical activity, dietary intakes). RESULTS: Our cohort included 166 (4.5%) short sleepers and 505 (14%) long sleepers. Shorter sleep duration was more likely to be observed in women aged ≥35 years who were multiparous, and had previous pregnancy, insufficient gestational weight gain, engaged in more vigorous physical activity, drank alcohol, were vegan and/or never took folic-acid supplements. Compared with normal sleepers (29%), the prevalence of GDM was significantly higher in short sleepers (38%; P = 0.01), but not in long sleepers (31%; P = 0.224). In the multivariable model, women with short sleep durations during early pregnancy had a 32% greater risk of GDM [adjusted risk ratio (aRR): 1.32, 95% CI: 1.06-1.63], whereas long sleepers did not (aRR: 1.09, 95% CI: 0.94-1.26). CONCLUSION: Short sleep duration during early pregnancy is associated with an increased risk of GDM. This suggests that more attention should be paid to controlling the development of GDM in pregnant women with insufficient sleep.

Exploring the protective effect of ShengMai-Yin and Ganmaidazao decoction combination against type 2 diabetes mellitus with nonalcoholic fatty liver disease by network pharmacology and validation in KKAy mice.

ETHNOPHARMACOLOGICAL RELEVANCE: ShengMai-Yin and Ganmaidazao decoction are classic formulas in traditional Chinese medicine. Individually, Shengmai-Yin is used to treat cardiovascular diseases, and Ganmaidazao decoction for therapy of mental disorders. The combination of Shengmai-Yin and Ganmaidazao decoction (SGD) is normally used as adjuvant therapy for type 2 diabetes mellitus (T2DM). AIM OF THE STUDY: The central aim is to elucidate the pharmacological efficacy of SGD and its mechanism in the treatment of T2DM with non-alcoholic fatty liver disease (NAFLD). MATERIALS AND METHODS: Active ingredients in SGD and their drug targets were identified using network analysis followed by experimental validation. First, existing databases were mined for information relevant to SGD, including pharmacological actions, chemical components, physicochemical characteristics, potential targets, and implicated diseases. Candidate patterns obtained with the network analysis were then tested in a KKAy mouse model of T2DM with NAFLD. Various doses of SGD were administered, followed by measurements of fasting blood glucose, oral glucose tolerance tests, insulin tolerance tests, markers of lipid metabolism - including free fatty acids (FFA), triglycerides (TG), and total cholesterol (TC) - liver histology, and expression levels of implicated molecules including PI3K/AKT and PPARα. RESULTS: Over 300 potential active compounds with their physicochemical characteristics and 562 candidate targets were collected, and then the network of them was constructed. Follow-up pathway and functional enrichment analyses indicated that SGD influences metabolism-related signaling pathways including PI3K-Akt, AMPK, and PPAR. In validation experiments, treatment of KKAy mice with SGD reduced serum levels of glucose, TC, TG, and FFA, decreased numbers of crown-like structures in visceral adipose tissue, reduced adipocyte size, and lowered liver lipid deposits. Further, SGD improved liver metabolism by increasing the expressions of PPARα, HSL, and PI3K/Akt, and decreasing expressions of SREBP-1 and FASN, inhibiting lipid biosynthesis, and increasing insulin sensitivity. CONCLUSION: Experimental validation of network analysis revealed anti-diabetic effects of the plant product SGD, manifested most notably by improved serum profiles and diminished insulin resistance. These experimental results may have clinical implications.

Effects of environmentally relevant concentrations of tris (2-butoxyethyl) phosphate on growth and transcription of genes involved in the GH/IGF and HPT axes in zebrafish (Danio rerio).

Tris (2-butoxyethyl) phosphate (TBOEP), as one of the most widely used organophosphate flame retardants (OPFRs), is applied in nearly all manufactured items and materials. It has been reported that TBOEP could cause developmental impairments and disrupt the endocrine regulation of fish growth during acute toxic experiments. However, concentrations to which fish were exposed in these studies were greater than environmentally relevant concentrations ever reported. This study examined effects on growth associated with exposure of zebrafish to 0, 0.1, 1 and 10 µg/L TBOEP during 20-90 days post fertilization (dpf). The changes in growth indicators and bioaccumulation of TBOEP were examined along with the transcription of related genes in the growth hormone/insulin-like growth factor (GH/IGF) axis and the hypothalamic-pituitary-thyroid (HPT) axis. The average body contents of TBOEP were higher in females than in males in all the exposure groups. Exposure to environmentally relevant concentrations of TBOEP significantly decreased body length and body mass and down-regulated expression of several genes involved in the GH/IGF and HPT axes. Exposure to TBOEP decreased plasma thyroxine (T4) content accompanied by decreased mRNA level of thyrotropin ß-subunit (tshß) in females at 60 dpf, but no effects were observed at 90 dpf. These results suggested that bioaccumulation of TBOEP and down-regulation of genes involved in the GH/IGF axis might be responsible for the observed growth inhibition in zebrafish exposed to TBOEP.

Socioeconomic inequality in periconceptional folic acid supplementation in China: a census of 0.9 million women in their first trimester of pregnancy.

BACKGROUND: To assess socioeconomic inequality in periconceptional folic acid supplementation in China. METHODS: We used data of periconceptional folic acid (FA) supplementation of rural Chinese women from the National Free Preconception Health Examination Project from 2010 to 2012 and socioeconomic level data from the National Bureau of Statistics. We used logistic models to assess the associations between the prevalence of taking FA and the sociodemographic characteristics of the participants, the couples, and the socioeconomic levels of their region of residence. RESULTS: Of the 907,720 included women, 682,315 (75.62%) of the women reported taking FA. The prevalence of FA supplementation was significantly higher in participants aged 21-29 (75.87%) than in those women aged 40-49 (68.44%, p < 0.01). The prevalence of FA supplementation was significantly higher in the region with the highest Per Capita Gross Regional Product than in the regions with lower Per Capita Gross Regional Product (aOR = 12.20 [95% CI:9.54-15.61]). The higher the per capita net income of farmer households in the region, the higher the prevalence of FA supplementation (aOR = 1.95 [95% CI:1.74-2.18]). CONCLUSIONS: The rate of periconceptional FA supplementation among rural Chinese women has increased with the support of China's Health System Reform policy. However, socioeconomic disparities in periconceptional folic acid supplementation remain.

Hydrothermal Preparation, Crystal Chemistry, and Redox Properties of Iron Muscovite Clay.

The development of functional materials based on Earth-abundant, environmentally benign compositions is critical for ensuring their commercial viability and sustainable production. Here we present an investigation into the crystal chemistry and electrochemical properties of the muscovite clay KFe2.75Si3.25O10(OH)2. We first report a low-temperature hydrothermal reaction that allows for a significant degree of control over sample crystallinity, particle morphology, and cation distribution through the lattice. A complex sequence of stacking faults is identified and characterized using a combination of Mössbauer spectroscopy and total scattering neutron experiments. We then show the existence of a reversible electrochemical process using galvanostatic cycling with complementary cyclic voltammetry suggesting that the redox activity occurs primarily on the surface of the particles. We conclude by determining that the ability to (de)intercalate Li ions from the material is hindered by the strong negative charge on the transition metal silicate layers, which prevents the displacement of the interlayer K ions. This work calls attention to a hugely Earth-abundant family of minerals that possesses useful electrochemical properties that warrant further exploration.

Immune and Metabolic Regulation Mechanism of Dangguiliuhuang Decoction against Insulin Resistance and Hepatic Steatosis.

Dangguiliuhuang decoction (DGLHD) is a traditional Chinese medicine (TCM) formula, which mainly consists of angelica, radix rehmanniae, radix rehmanniae praeparata, scutellaria baicalensis, coptis chinensis, astragalus membranaceus, and golden cypress, and used for the treatment of diabetes and some autoimmune diseases. In this study, we explored the potential mechanism of DGLHD against insulin resistance and fatty liver in vivo and in vitro. Our data revealed that DGLHD normalized glucose and insulin level, increased the expression of adiponectin, diminished fat accumulation and lipogenesis, and promoted glucose uptake. Metabolomic analysis also demonstrated that DGLHD decreased isoleucine, adenosine, and cholesterol, increased glutamine levels in liver and visceral adipose tissue (VAT) of ob/ob mice. Importantly, DGLHD promoted the shift of pro-inflammatory to anti-inflammatory cytokines, suppressed T lymphocytes proliferation, and enhanced regulatory T cells (Tregs) differentiation. DGLHD also inhibited dendritic cells (DCs) maturation, attenuated DCs-stimulated T cells proliferation and secretion of IL-12p70 cytokine from DCs, and promoted the interaction of DCs with Tregs. Further studies indicated that the changed PI3K/Akt signaling pathway and elevated PPAR-γ expression were not only observed with the ameliorated glucose and lipid metabolism in adipocytes and hepatocytes, but also exhibited in DCs and T cells by DGLHD. Collectively, our results suggest that DGLHD exerts anti-insulin resistant and antisteatotic effects by improving abnormal immune and metabolic homeostasis. And DGLHD may be a novel approach to the treatment of obesity-related insulin resistance and hepatic steatosis.

[Comparative study on therapeutic effect between acupuncture at special acupoints and non-specific acupoints in foot yangming meridian for functional dyspepsia].

OBJECTIVE: To compare the clinical efficacy differences among acupuncture at special acupoints, nonspecific acupoints and non acupoints in Foot Yangming Meridian for functional dyspepsia (FD) at different time points. METHODS: One hundred and sixteen FD patients were randomly divided into a special acupoints in Foot Yangming Meridian group (group A, n = 36), a non-specific acupoints in Foot Yangming Meridian group (group B, n 39) and a non acupoints group (group C, n = 41). Group A was treated with acupuncture at Chongyang (ST 42), Fenglong (ST 40), Zusanli (ST 36) and Liangqiu (ST 34), and group B was treated with acupuncture at Tiaokou (ST 38), Yinshi (ST 33), Futu (ST 32) and Dubi (ST 35), and the assisted acupoints were setting up at 2 mm apart from selected acupoints in meridian of proximal part. Group C was treated with 4 non acupoints: (1) medial elbow, middle point on the line between olecranon and armpit; (2) middle point on the line between condylus medialis humeri and wrist of elbow-bone; (3) the junction of deltoid and biceps inside the arm; (4) 1-2 cm horizontally away from Zusanli (ST 36), lateral border of shin bone, and the assisted acupoints were setting up at 2 mm away from selected non acupoints along limb longitudinal axis of proximal part. All acupoints and assisted acupoints were connected to the HANS nerve stimulator and they were all treated for 4 courses. The FD Symptom Index (FDI) and MOS 36-Item Short-Form Health Survey (SF-36) after treatment, 1 month and 3 months after treatment were observed and the clinical efficacy was assessed. RESULTS: After treatment, the total effective rate of abdominal fullness after meal, early satiation, upper abdominal pain and upper abdominal burning sensation were 84.8% (28/33), 67.7% (21/31), 76.9% (20/26) and 56.3% (9/16), respectively, in group A, and 45.9% (17/37), 38.7% (12/31), 42.9% (12/28) and 38.5% (5/13), respectively, in group B, and 15.8% (6/38), 18.4% (7/38), 46.1% (12/26) and 16.7% (4/24), respectively, in group C. The total effective rate of abdominal fullness after meal, early satiation and upper abdominal pain in group A were superior to those in group B (all P < 0.05), and the total effective rate of abdominal fullness after meal, early satiation, upper abdominal pain and upper abdominal burning sensation in group A were all superior to those in group C (all P < 0.05), and the total effective rate of abdominal fullness after meal in group B was superior to that in group C (P < 0.05). The score of FDI and SF-36 after treatment, 1 month and 3 months after treatment in all the groups were better than those before treatment (all P < 0.05), and above indices in group A were the most significant (all P < 0.05), and the scores of FDI and SF-36 after treatment in group B were better than those in group C (both P < 0.05). CONCLUSION: All the treatment of acupuncture at special acupoints, non-specific acupoints in Foot Yangming Meridian and non acupoints have therapeutic effect on FD, but acupuncture at special acupoints has better short and long term therapeutic effects, which confirm the existence of acupoints specificity.

Galvanic zinc-copper microparticles produce electrical stimulation that reduces the inflammatory and immune responses in skin.

The human body has its own innate electrical system that regulates the body's functions via communications among organs through the well-known neural system. While the effect of low-level electrical stimulation on wound repair has been reported, few studies have examined the effect of electric potential on non-wounded, intact skin. A galvanic couple comprised of elemental zinc and copper was used to determine the effects of low-level electrical stimulation on intact skin physiology using a Dermacorder device. Zn-Cu induced the electrical potential recorded on intact skin, enhanced H(2)O(2) production and activated p38 MAPK and Hsp27 in primary keratinocytes. Treatment with Zn-Cu was also found to reduce pro-inflammatory cytokines, such as IL-1α, IL-2, NO and TNF-α in multiple cell types after stimulation with PHA or Propionibacterium acnes bacteria. The Zn-Cu complex led to a dose-dependent inhibition of TNF-α-induced NF-κB levels in keratinocytes as measured by a dual-luciferase promoter assay, and prevented p65 translocation to the nucleus observed via immunofluorescence. Suppression of NF-κB activity via crosstalk with p38 MAPK might be one of the potential pathways by which Zn-Cu exerted its inflammatory effects. Topical application of Zn-Cu successfully mitigated TPA-induced dermatitis and oxazolone-induced hypersensitivity in mice models of ear edema. Anti-inflammatory activity induced by the Zn-Cu galvanic couple appears to be mediated, at least in part, by production of low level of hydrogen peroxide since this activity is reversed by the addition of Catalase enzyme. Collectively, these results show that a galvanic couple containing Zn-Cu strongly reduces the inflammatory and immune responses in intact skin, providing evidence for the role of electric stimulation in non-wounded skin.

Activation of phosphatidylinositol-linked novel D1 dopamine receptor contributes to the calcium mobilization in cultured rat prefrontal cortical astrocytes.

Recent evidences indicate the existence of an atypical D(1) dopamine receptor other than traditional D(1) dopamine receptor in the brain that mediates PI hydrolysis via activation of phospholipase C(beta) (PLC(beta)). To further understand the basic physiological function of this receptor in brain, the effects of a selective phosphoinositide (PI)-linked D(1) dopamine receptor agonist SKF83959 on cytosolic free calcium concentration ([Ca(2+)](i)) in cultured rat prefrontal cortical astrocytes were investigated by calcium imaging. The results indicated that SKF83959 caused a transient dose-dependent increase in [Ca(2+)](i). Application of D(1) receptor, but not D(2), alpha(1) adrenergic, 5-HT receptor, or cholinergic antagonist prevented SKF83959-induced [Ca(2+)](i) rise, indicating that activation of the D(1) dopamine receptor was essential for this response. Increase in [Ca(2+)](i) was a two-step process characterized by an initial increase in [Ca(2+)](i) mediated by release from intracellular stores, supplemented by influx through voltage-gated calcium channels, receptor-operated calcium channels, and capacitative Ca(2+) entry. Furthermore, SKF83959-stimulated increase in [Ca(2+)](i) was abolished following treatment with a PLC inhibitor. Overall, these results suggested that activation of D(1) receptor by SKF83959 mediates a dose-dependent mobilization of [Ca(2+)](i) via the PLC signaling pathway in cultured rat prefrontal cortical astrocytes.

Puerarin facilitates Ca(2+)-induced Ca(2+) release triggered by KCl-depolarization in primary cultured rat hippocampal neurons.

The effects of puerarin on behaviour and brain neuronal activity in animal studies have been described previously. However, molecule mechanisms underlying these effects were poorly understood. Here, we examined the regulation of puerarin on the Ca(2+) signals in primary rat hippocampal neurons using Fura-2 based calcium imaging techniques. Application of puerarin had no effect on the basal intracellular calcium concentration ([Ca(2+)](i)), but potentiated the KCl-evoked [Ca(2+)](i) transient in 87% of recorded neurons. Dantrolene or ruthenium red, the inhibitors of ryanodine receptors, completely blocked this potentiation induced by puerarin. Moreover, in Ca(2+)-free solution, pre-application of puerarin significantly augmented the elevation of [Ca(2+)](i) evoked by caffeine (3 mM), which is a specific agent to activate the ryanodine receptors. In contrast, nifedipine failed to prevent the potentiation induced by puerarin. Similarly, in the experiments of whole-cell patch-clamp recording, puerarin did not show any effect on calcium currents generated by depolarization pulses. These data demonstrated that the potentiation induced by puerarin was attributed to the facilitation of Ca(2+)-induced Ca(2+) release (CICR) via ryanodine receptors, rather than extracellular Ca(2+) influx. Using estrogen receptor antagonist ICI 182780 and tamoxifen, we further demonstrated that the potentiation induced by puerarin was mediated by the estrogen receptor. Furthermore, the membrane-permeant inhibitor of protein kinase A (PKA) H89 completely inhibited this potentiation. However, U-73122, the inhibitor of phospholipase C (PLC) had no effect, indicating that the cyclic AMP/PKA signaling pathway was involved in the activation of CICR by puerarin.