Polycystin-2 is a novel cation channel implicated in defective intracellular Ca(2+) homeostasis in polycystic kidney disease.

Mutations in polycystins-1 and -2 (PC1 and PC2) cause autosomal dominant polycystic kidney disease (ADPKD), which is characterized by progressive development of epithelial renal cysts, ultimately leading to renal failure. The functions of these polycystins remain elusive. Here we show that PC2 is a Ca(2+)-permeable cation channel with properties distinct from any known intracellular channels. Its kinetic behavior is characterized by frequent transitions between closed and open states over a wide voltage range. The activity of the PC2 channel is transiently increased by elevating cytosolic Ca(2+). Given the predominant endoplasmic reticulum (ER) location of PC2 and its unresponsiveness to the known modulators of mediating Ca(2+) release from the ER, inositol-trisphosphate (IP(3)) and ryanodine, these results suggest that PC2 represents a novel type of channel with properties distinct from those of the other Ca(2+)-release channels. Our data also show that the PC2 channel can be translocated to the plasma membranes by defined chemical chaperones and proteasome modulators, suggesting that in vivo, it may also function in the plasma membrane under specific conditions. The sensitivity of the PC2 channel to changes of intracellular Ca(2+) concentration is deficient in a mutant found in ADPKD patients. The dysfunction of such mutants may result in defective coupling of PC2 to intracellular Ca(2+) homeostasis associated with the pathogenesis of ADPKD.

[Screening of anti-bacteria activity of extracts of Gaultheria leucocarpa var. yunnanensis].

OBJECTIVE: To further develop Gaultheria leucocarpa var. yunnanensis, anti-bacteria constituents in it were screened. METHOD: The constituents were extracted by chromatographic process. The anti-bacteria test was made with regulatory method of analysis. RESULT AND CONCLUSION: Anti-bacteria test with extracts of water, acetic ester and n-butanol showed that 3 extracts from 22 samples had anti-Staphylococcus aureus action, and the extracts from root and stem showed the same result. 2 extracts could kill Escherichia coli and Pseudomonas aeruginosa. The lower the concentrate, the less the anti-bacteria action was. These results suggested that not only essential oil but other ingredients from G. leucocarpa var.yunnanensis have anti-bacteria activity. Anti-fungi test of the same extracts didn't indicate remarkable action.

[Studies on chemical constituents of Gaultheria leucocarpa var. Yunnanensis (Franch.) T. Z. Hsu & R. C. Fang].

OBJECTIVE: To separate and identify the chemical constituents of the aerial part of Gaultheria leucocarpa var. yunnanensis. METHOD: The compounds were extracted with solvents, isolated by column chromatography and identified by spectral analysis. RESULT: Four compounds were identified as n-dotriacontane and its homologous compound(1), ursolic acid(2), vanillic acid(3), and quercitrin(4). CONCLUSION: The compounds 1, 4 were obtained from the plant for the first time, and 2 and 3 were from above-ground part of the plant for the first time.

[Studies on resources of ethnomedicine Gaultheria leucocarpa var. yunnanensis].

OBJECTIVE: To study the resources of G. leucocarpa var. yunnanensis for further development of this drug. METHOD: Field investigating, consulting with relevant experts and looking into available specimens. RESULTS AND CONCLUSION: G. leucocarpa var. yunnanensis is widely distributed in the southern regions of the Yangtze River. The field investigation suggests that the distribution center is situated in Yunnan province, mainly in Kunming, Chuxiong and Dali counties. The climate in these areas is moderate and moist. G. leucocarpa var. yunnanensis is not a dominant species in this natural environment. In some places, it grows with other species of Gaultheria, such as G. fragrantissima, G. tetramera, G. griffithiana and G. leucocarpa var. cumingiana. It is distributed at altitudes from 400 m to 3,500 m. Accustomed to different sunshine conditions, G. leucocarpa var. yunnanensis prefers stronger sunlight and commonly grows on sunny slopes, seldom in dense forest, propagating itself by roots. As a folk medicine, G. leucocarpa var. yunnanensis is commonly used to treat rheumatic arthritis(RA), dazzling, suppressed menstruation, cold, cough, asthma, strain hematemesis, eczema, ascites, wound, amebic dysentery, acute and chronic prostatitis. It is suggested that further pharmacological and clinical researches of this plant be concentrated on the treatment of RA and relief of aches.

A rat kidney-specific calcium transporter in the distal nephron.

Active absorption of calcium from the intestine and reabsorption of calcium from the kidney are major determinants of whole body calcium homeostasis. Two recently cloned proteins, CaT1 and ECaC, have been postulated to mediate apical calcium uptake by rat intestine and rabbit kidney, respectively. By screening a rat kidney cortex library with a CaT1 probe, we isolated a cDNA encoding a protein (CaT2) with 84.2 and 73.4% amino acid identities to ECaC and CaT1, respectively. Unlike ECaC, CaT2 is kidney-specific in the rat and was not detected in intestine, brain, adrenal gland, heart, skeletal muscle, liver, lung, spleen, thymus, and testis by Northern analysis or reverse transcription polymerase chain reaction. The expression pattern of CaT2 in kidney was similar to that of calbindin D(28K) and the sodium calcium exchanger 1, NCX1, by in situ hybridization of adjacent sections. Furthermore, the mRNAs for CaT2 and calbindin D(28K) were colocalized in the same cells. CaT2 mediated saturable calcium uptake with a Michaelis constant (K(m)) of 0.66 mm when expressed in Xenopus laevis oocytes. Under voltage clamp condition, CaT2 promoted inward currents in X. laevis oocytes upon external application of Ca(2+). Sr(2+) and Ba(2+) but not Mg(2+) also evoked inward currents in CaT2-expressing oocytes. Similar to the alkaline earth metal ions, application of Cd(2+) elicited inward current in CaT2-expressing oocytes with a K(m) of 1.3 mm. Cd(2+), however, also potently inhibited CaT2-mediated Ca(2+) uptake with an IC(50) of 5.4 micrometer. Ca(2+) evoked currents were reduced at low pH and increased at high pH and were only slightly affected by the L-type voltage-dependent calcium channel antagonists, nifedipine, verapamil, diltiazem, and the agonist, Bay K 8644, even at relatively high concentrations. In conclusion, CaT2 may participate in calcium entry into the cells of the distal convoluted tubule and connecting segment of the nephron, where active reabsorption of calcium takes place via the transcellular route. The high sensitivity of CaT2 to Cd(2+) also provides a potential explanation for Cd(2+)-induced hypercalciuria and resultant renal stone formation.

Yeast SMF1 mediates H(+)-coupled iron uptake with concomitant uncoupled cation currents.

Yeast membrane proteins SMF1, SMF2, and SMF3 are homologues of the DCT1 metal ion transporter family. Their functional characteristics and the implications of these characteristics in vivo have not yet been reported. Here we show that SMF1 expressed in Xenopus oocytes mediates H(+)-dependent Fe(2+) transport and uncoupled Na(+) flux. SMF1-mediated Fe(2+) transport exhibited saturation kinetics (K(m) = 2.2 microM), whereas the Na(+) flux did not, although both processes were electrogenic. SMF1 is also permeable to Li(+), Rb(+), K(+), and Ca(2+), which likely share the same uncoupled pathway. SMF2 (but not SMF3) mediated significant increases in both Fe(2+) and Na(+) transport compared with control oocytes. These data are consistent with the concept that uptake of divalent metal ions by SMF1 and SMF2 is essential to yeast cell growth. Na(+) inhibited metal ion uptake mediated by SMF1 and SMF2 expressed in oocytes. Consistent with this, we found that increased sensitivity of yeast to EGTA in the high Na(+) medium is due to inhibition of SMF1- and SMF2-mediated metal ion transport by uncoupled Na(+) pathway. Interestingly, DCT1 also mediates Fe(2+)-activated uncoupled currents. We propose that uncoupled ion permeabilities in metal ion transporters protect cells from metal ion overload.

Molecular cloning and characterization of a channel-like transporter mediating intestinal calcium absorption.

Calcium is a major component of the mineral phase of bone and serves as a key intracellular second messenger. Postnatally, all bodily calcium must be absorbed from the diet through the intestine. Here we report the properties of a calcium transport protein (CaT1) cloned from rat duodenum using an expression cloning strategy in Xenopus laevis oocytes, which likely plays a key role in the intestinal uptake of calcium. CaT1 shows homology (75% amino acid sequence identity) to the apical calcium channel ECaC recently cloned from vitamin D-responsive cells of rabbit kidney and is structurally related to the capsaicin receptor and the TRP family of ion channels. Based on Northern analysis of rat tissues, a 3-kilobase CaT1 transcript is present in rat duodenum, proximal jejunum, cecum, and colon, and a 6.5-kilobase transcript is present in brain, thymus, and adrenal gland. In situ hybridization revealed strong CaT1 mRNA expression in enterocytes of duodenum, proximal jejunum, and cecum. No signals were detected in kidney, heart, liver, lung, spleen, and skeletal muscle. When expressed in Xenopus oocytes, CaT1 mediates saturable Ca(2+) uptake with a Michaelis constant of 0.44 mM. Transport of Ca(2+) by CaT1 is electrogenic, voltage-dependent, and exhibits a charge/Ca(2+) uptake ratio close to 2:1, indicating that CaT1-mediated Ca(2+) influx is not coupled to other ions. CaT1 activity is pH-sensitive, exhibiting significant inhibition by low pH. CaT1 is also permeant to Sr(2+) and Ba(2+) (but not Mg(2+)), although the currents evoked by Sr(2+) and Ba(2+) are much smaller than those evoked by Ca(2+). The trivalent cations Gd(3+) and La(3+) and the divalent cations Cu(2+), Pb(2+), Cd(2+), Co(2+), and Ni(2+) (each at 100 microM) do not evoke currents themselves, but inhibit CaT1-mediated Ca(2+) transport. Fe(3+), Fe(2+), Mn(2+), and Zn(2+) have no significant effects at 100 microM on CaT1-mediated Ca(2+) transport. CaT1 mRNA levels are not responsive to 1,25-dihydroxyvitamin D(3) administration or to calcium deficiency. Our studies strongly suggest that CaT1 provides the principal mechanism for Ca(2+) entry into enterocytes as part of the transcellular pathway of calcium absorption in the intestine.

A family of mammalian Na+-dependent L-ascorbic acid transporters.

Vitamin C (L-ascorbic acid) is essential for many enzymatic reactions, in which it serves to maintain prosthetic metal ions in their reduced forms (for example, Fe2+, Cu+), and for scavenging free radicals in order to protect tissues from oxidative damage. The facilitative sugar transporters of the GLUT type can transport the oxidized form of the vitamin, dehydroascorbic acid, but these transporters are unlikely to allow significant physiological amounts of vitamin C to be taken up in the presence of normal glucose concentrations, because the vitamin is present in plasma essentially only in its reduced form. Here we describe the isolation of two L-ascorbic acid transporters, SVCT1 and SVCT2, from rat complementary DNA libraries, as the first step in investigating the importance of L-ascorbic acid transport in regulating the supply and metabolism of vitamin C. We find that SVCT1 and SVCT2 each mediate concentrative, high-affinity L-ascorbic acid transport that is stereospecific and is driven by the Na+ electrochemical gradient. Despite their close sequence homology and similar functions, the two isoforms of the transporter are discretely distributed: SVCT1 is mainly confined to epithelial systems (intestine, kidney, liver), whereas SVCT2 serves a host of metabolically active cells and specialized tissues in the brain, eye and other organs.

[Clinical and experimental study on rapid bladder ultrasound developer of Chinese medicinal herbs].

OBJECTIVE: To seek for a rapid type B ultrasound developer of Chinese medicinal herbs, so that the bladder and pelvic cavity developed clearly and pelvic cavity diseases could be diagnosed rapidly. METHODS: One hundred and twenty-two patients were observed clinically and animal experiments were performed. The rapid bladder ultrasonography developer (RBUD-1, a preparation of Chinese herbal medicine) alone was used in Group 1, composite prescription of Western and Chinese medicine was used in group 2. The control groups were using lasix or mineral water. RESULTS: Rapid diuresis and the decrease of the bladder capacity needed for development could be realized by Chinese medical herbs preparation, the difference between Group 1 and control group in developing time and bladder capacity were very significant. Results of animal experiments, which were referred to clinical grouping, showed the diuretic intensity of RBUD-1 within one hour was significantly higher than that in the other groups. Toxicological study showed the RBUD-1 was a non-toxic preparation. CONCLUSION: RBUD-1 could effectively develop bladder and pelvic cavity, it would help to diagnose in time, on the other hand, it would also contribute for the combination imaging of Chinese and Western medicine.

[Quantitative analysis of synephrine in zhishi injection by RP-HPLC].

A reversed phase HPLC method was developed for the determination of synephrine in Zhishi Injection using methanol-water (1:1) containing sodium 1-pentane sulfonate (3.5%, G/V) and acetic acid (0.1%, V/V) as the mobile phase with UV detoction at 275nm. Linear response was obtained in the range of 8-64 micrograms/ml (r = 0.9998), the absolute recovery was 96.2-103.4%, and RSD 2.7%-6.0% (n = 7).

[A study on combined acupuncture and enflurane anesthesia for craniotomy].

The reinforcing effect of transcutaneous acupoint electric stimulation (TAES) with enflurane anesthesia during craniotomy was studied. 110 neurosurgical patients were randomly divided into three groups. Anesthesia was maintained with enflurane in group A (n = 40); in group B, enflurane anesthesia was supplemented by TAES with Han's acupoint nerve stimulator (HANS) at Hegu, Yuyao and Fengchi points on the operated side (n = 40); in group C, enflurane anesthesia was supplemented by TAES and scalp infiltration with 0.5% procaine (n = 30). The results showed that the minimum alveolar concentration (MAC) of enflurane in group B and C decreased 37.8-47.0% and 42.1-66.1% respectively than that in group A. The hemodynamics was more stable during operation, and the patients recovered faster after operation in group B and C. It was concluded that TAES with HANS significantly potentiated the anesthetic effect and decreased the side effects of enflurane during operation, and that the triple combination of TAES, enflurane and scalp infiltration with procaine proved to be a better anesthetic method for craniotomy.