Antibacterial property of isoflavonoids isolated from Erythrina variegata against cariogenic oral bacteria.

The antibacterial property of 7 compounds, isolated from Erythrina variegata (Leguminosae) by repeated silica gel column chromatography, against cariogenic oral bacteria was investigated. Extensive spectroscopic study revealed that all were isoflavonoids. Among them, 3,9-dihydroxy-2,10-di(gamma,gamma-dimethylallyl)-6a,11a-dehydropterocarpan (erycristagallin) showed the highest antibacterial activity against mutans streptococci, other oral streptococci, Actinomyces and Lactobacillus species with a minimum inhibitory concentration (MIC) range of 1.56-6.25 microg/ml, followed by 3,6a-dihydroxy-9-methoxy-2,10-di(gamma,gamma-dimethylallyl)pterocarpan (erystagallinA) and 9-hydroxy-3-methoxy-2-gamma,gamma-dimethylallylpterocarpan (orientanol B) (MIC range: 3.13-12.5 microg/ml). The antibacterial effect of erycristagallin to mutans streptococci was based on a bactericidal action. Erycristagallin (6.25 microg/ml: MIC) completely inhibited incorporation of radio-labelled thymidine into Streptococcus mutans cells. Incorporation of radio-labelled glucose into bacterial cells was also strongly inhibited at MIC, and 1/2 MIC of the compound reduced the incorporation approximately by half. The findings indicate that erycristagallin has a potential as potent phytochemical agent for prevention of dental caries by inhibiting the growth of cariogenic bacteria and by interfering with incorporation of glucose responsible for production of organic acids.

Endotoxin contamination in wound dressings made of natural biomaterials.

Contamination by endotoxin of nine kinds of wound dressings made of natural biomaterials (calcium alginate, collagen, chitin, and poly-L-leucine) was examined with the use of water extracts. By applying the Limulus amoebocyte lysate (LAL) test, high concentrations of endotoxin were detected in extracts from three kinds of products made of calcium alginate. These extracts evoked fever in rabbits and induced the release of a proinflammatory (pyrogenic) cytokine, interleukin-6 (IL-6), from human monocytic cells (MM6-CA8). The effects disappeared when the extracts were treated with endotoxin-removing gel column chromatography or with an endotoxin antagonist, B464, confirming that the contaminating pyrogen was endotoxin. A noteworthy finding was that one of the endotoxin-containing extracts showed very weak IL-6-inducibility in human monocytic cells in contrast to its high pyrogenicity to rabbits. The discrepancy could be explained based on differences between humans and rabbits in sensitivity to the endotoxin, because the extract showed higher proinflammatory-cytokine (TNF-alpha)-inducibility in rabbit whole-blood cells (WBCs) than human WBCs. The results suggest that the LAL test is a useful method of detecting endotoxin contamination in wound dressings and the MM6-CA8 assay is a good supplement to the LAL test for evaluating pyrogenicity in humans accurately.

Antibacterial activity of isoflavonoids isolated from Erythrina variegata against methicillin-resistant Staphylococcus aureus.

AIMS: To screen 16 isoflavonoids isolated from Erythrina variegata (Leguminosae) for their antibacterial activity against methicillin-resistant Staphylococcus aureus (MRSA). METHODS AND RESULTS: The roots of E. variegata were macerated with acetone. The chloroform-soluble fraction of the residue was subjected to repeated silica gel column chromatography followed by elution with various solvents. Structures of the isolated compounds were determined by extensive spectroscopic studies. Each compound was dissolved in dimethyl sulphoxide and added to agar plates (final concentration 1.56-100 microg ml(-1) and suspensions of MRSA spotted onto the agar plates to determine the minimum inhibitory concentration (MIC). Repeated silica gel chromatography yielded 16 compounds and spectroscopic studies revealed that all were isoflavonoids. Whilst 14 compounds showed antibacterial activity in this concentration range, the MIC values varied significantly among them. Of the active compounds, 3,9-dihydroxy-2,10-di(gamma,gamma-dimethylallyl)-6a,11a-dehydropterocarpan (erycristagallin) and 9-hydroxy-3-methoxy-2-gamma,gamma-dimethylallylpterocarpan (orientanol B) exhibited the highest activity with MIC values of 3.13-6.25 microg ml(-1). CONCLUSIONS: Erycristagallin and orientanol B showed the highest anti-MRSA activity (3.13-6.25 microg ml(-1). SIGNIFICANCE AND IMPACT OF THE STUDY: Erycristagallin and orientanol B could be leading compounds for phytotherapeutic agents against MRSA infections.

Anti-angiogenic action of hyperthermia by suppressing gene expression and production of tumour-derived vascular endothelial growth factor in vivo and in vitro.

Vascular endothelial growth factor is an important angiogenic factor for tumour progression because it increases endothelial-cell proliferation and remodels extracellular matrix in blood vessels. We demonstrated that hyperthermia at 42 degrees C, termed heat shock, suppressed the gene expression and production of vascular endothelial growth factor in human fibrosarcoma HT-1080 cells and inhibited its in vitro angiogenic action on human umbilical vein endothelial cells. The gene expression of alternative splicing variants for vascular endothelial growth factor, VEGF121, VEGF165 and VEGF189, was constitutively detected in HT-1080 cells, but the VEGF189 transcript was less abundant than VEGF121 and VEGF165. When HT-1080 cells were treated with heat shock at 42 degrees C for 4 h and then maintained at 37 degrees C for another 24 h, the gene expression of all vascular endothelial growth factor variants was suppressed. In addition, HT-1080 cells were found to produce abundant VEGF165, but much less VEGF121, both of which were inhibited by heat shock. Furthermore, the level of vascular endothelial growth factor in sera from six cancer patients was significantly diminished 2-3 weeks after completion of whole-body hyperthermia at 42 degrees C (49.9+/-36.5 pg x ml(-1), P<0.01) as compared with that prior to the treatment (177.0+/-77.5 pg x ml(-1)). On the other hand, HT-1080 cell-conditioned medium showed vascular endothelial growth factor-dependent cell proliferative activity and the augmentation of pro-matrix metalloproteinase-1 production in human umbilical vein endothelial cells. The augmentation of endothelial-cell proliferation and pro-matrix metalloproteinase-1 production was poor when human umbilical vein endothelial cells were treated with conditioned medium from heat-shocked HT-1080 cells. These results suggest that hyperthermia acts as an anti-angiogenic strategy by suppressing the expression of tumour-derived vascular endothelial growth factor production and thereby inhibiting endothelial-cell proliferation and extracellular matrix remodelling in blood vessels.

Revised structures for senegalensin and euchrenone b(10).

Two prenylated isoflavones (1 and 2) with a hydroxyisopropyldihydrofuran moiety have been isolated from the wood of Erythrina suberosa var. glabrescence. The structure of compound 1 was in agreement with that of the previously reported senegalensin, isolated from the stem bark of Erythrina senegalensis. The structure of senegalensin was revised from structure 2 to structure 1 by spectroscopic means. Compound 2, the regioisomer of 1, was confirmed as euchrenone b(10) by comparison with the spectral data of the reported euchrenone b(10), isolated from the roots of Euchresta horsfieldii. The structure of 2 was established by 2D NMR spectroscopic analysis and by the X-ray crystallographic analysis of its p-bromobenzoyl derivative (2b).

Cathelicidin family of antibacterial peptides CAP18 and CAP11 inhibit the expression of TNF-alpha by blocking the binding of LPS to CD14(+) cells.

Mammalian myeloid and epithelial cells express several kinds of antibacterial peptides (alpha-/beta-defensins and cathelicidins) that contribute to the innate host defense by killing invading micro-organisms. In this study we evaluated the LPS-neutralizing activities of cathelicidin peptides human CAP18 (cationic antibacterial proteins of 18 kDa) and guinea pig CAP11 using the CD14(+) murine macrophage cell line RAW264.7 and the murine endotoxin shock model. Flow cytometric analysis revealed that CAP18 and CAP11 inhibited the binding of FITC-conjugated LPS to RAW264.7 cells. Likewise, Northern and Western blot analyses indicated that CAP18 and CAP11 suppressed LPS-induced TNF-alpha mRNA and protein expression by RAW264.7 cells. Interestingly, CAP18 and CAP11 possessed LPS-binding activities, and they strongly suppressed the interaction of LPS with LPS binding protein that mediates the transport of LPS to CD14 to facilitate the activation of CD14(+) cells by LPS. Moreover, when CAP18 and CAP11 were preincubated with RAW264.7 cells, they bound to the cell surface CD14 and inhibited the binding of FITC-LPS to the cells. Furthermore, in the murine endotoxin shock model, CAP18 or CAP11 administration inhibited the binding of LPS to CD14(+) cells (peritoneal macrophages) and suppressed LPS-induced TNF-alpha expression by these cells. Together these observations indicate that cathelicidin peptides CAP18 and CAP11 probably exert protective actions against endotoxin shock by blocking the binding of LPS to CD14(+) cells, thereby suppressing the production of cytokines by these cells via their potent binding activities for LPS and CD14.

Chemical and biological evaluation of endotoxin contamination on natural rubber latex products.

Relationship between pyrogenicity and bacterial endotoxin contamination on latex products was demonstrated by chemical analysis and biological assays. In commercially available latex products' surveillance, water extracts prepared from one surgical glove and two silicone elastomer-coated Foley catheters sterilized by gamma-irradiation were obviously pyrogenic in rabbits. The induced fever was monophasic at low dose of the pyrogenic extracts and biphasic at high dose. These extracts exhibited limulus amebocyte lysate gelation activity, and induced inflammatory cytokine (interleukin-1, interleukin-6, and tumor necrosis factor-alpha) production from MM6-CA8 human monocytoid cells. These biological properties, including pyrogenicity, completely disappeared by treating the pyrogenic extracts with endotoxin-adsorbent affinity column. Limulus amebocyte lysate activity and cytokine production from MM6-CA8 cells induced by the extracts were significantly decreased by endotoxin inhibitors, an active fragment peptide of an 18-kDa cationic antimicrobial protein and a synthetic lipid A B464 analogue. Furthermore, very small amounts of 2-keto-3-deoxyoctonate and 3-hydroxy fatty acid, which are common constituents of bacterial endotoxins, were detected by gas chromatography-mass spectrometry analysis of the pyrogenic extracts. These findings clearly showed that the pyrogenicity found in these latex products originated from endotoxins contaminating the products.

Motor cortex stimulation for deafferentation pain.

OBJECT: The authors tested a modified motor cortex stimulation (MCS) protocol for the treatment of deafferentation pain in 15 patients: eight patients with poststroke pain, four with brachial plexus injury, two with phantom limb pain, and one with spinal cord injury. METHODS: Preoperative pharmacological tests were performed with phentolamine, lidocaine, ketamine, thiopental, morphine, and a placebo. In 12 patients we placed a 20- or 40-grid electrode in the subdural space to determine the best stimulation point for pain relief over a few weeks and therefore the optimum position for a permanent internal device. In four patients, the MCS devices were implanted in the interhemispheric fissure to reduce lower-extremity pain. In one patient, the MCS device was placed within the central sulcus, and a 20-grid electrode was placed on the brain surface. In two patients with pain extending from the upper extremity to the hyperbody, dual-electrode devices were implanted to drive two electrodes. In 10 of the 15 patients MCS-induced pain reduction was achieved (four with excellent, two with good, and four with fair alleviation of pain). The result of pharmacological testing indicated that patients with ketamine sensitivity seem to be good candidates for MCS. CONCLUSIONS: Test stimulation with a subdural multigrid electrode was helpful in locating the best stimulation point for pain relief.

Motor cortex stimulation for central and peripheral deafferentation pain. Report of eight cases.

The authors tested a modified motor cortex stimulation protocol for treatment of central and peripheral types of deafferentation pain. Four patients with thalamic pain and four with peripheral deafferentation pain were studied. Preoperative pharmacological tests of pain relief were performed using phentolamine, lidocaine, ketamine, thiopental, and placebo. In five patients we placed a 20- or 40-electrode grid in the subdural space to determine the best stimulation point for pain relief for a few weeks before definitive placement of a four-electrode array. In three patients, the four-electrode array was implanted in the interhemispheric fissure as a one-stage procedure to treat lower-extremity pain. In two patients with pain extending from the extremity to the trunk or hip, dual devices were implanted to drive two electrodes. Six of eight patients experienced pain reduction (two each with excellent, good, and fair relief) from motor cortex stimulation. No correlation was apparent between pharmacological test results and the effectiveness of motor cortex stimulation. Patients with peripheral deafferentation pain, including two with phantom-limb pain and two with brachial plexus injury, attained pain relief from motor cortex stimulation, with excellent results in two cases. Testing performed with a subdural multiple-electrode grid was helpful in locating the best stimulation point for pain relief. Motor cortex stimulation may be effective for treating peripheral as well as central deafferentation pain.

Effect of dietary fat on lymphocyte proliferation and metabolism.

The effect of diets enriched with fat containing different fatty acids on glucose and glutamine metabolism of mesenteric lymph nodes lymphocytes, spleen, and thymus and lymphocyte proliferation was examined. The following fat-rich diets were tested: (1) standard chow (CC); (2) medium chain saturated fatty acids (MS)--coconut fat oil; (3) long chain saturated fatty acids (LS)--cocoa butter; (4) monounsaturated fatty acids (MU)--canola oil (n-9); (5) polyunsaturated fatty acids (PU)--soybean oil (n-6). Of the fat-rich diets tested, MS was the one to present the least pronounced effect. Lymphocyte proliferation was reduced by LS (64 per cent), MU (55 per cent), and PU (60 per cent). Hexokinase activity was enhanced in lymph node lymphocytes by PU (67 per cent), in the spleen by MS (42 per cent), and in the thymus by PU (30 per cent). This enzyme activity was reduced in the spleen (33 per cent) by LS and MU (35 per cent). In the thymus, this enzyme activity was reduced by LS (26 per cent) and MU (13 per cent). Maximal phosphate-dependent glutaminase activity was raised in lymphocytes by MS (70 per cent) and MU (20 per cent). This enzyme activity, however, was decreased in lymphocytes by PU (26 per cent), in the spleen by LS (15 per cent), and in the thymus by MU (44 per cent). Citrate synthase activity was increased in lymphocytes by MU (35 per cent), in the spleen by LS (56 per cent) and MU (68 per cent), and in the thymus by LS (42 per cent). This enzyme activity was decreased in lymphocytes by PU (24 per cent) only. [U-14C]-Glucose decarboxylation was raised by all fat-rich diets; MS (88 per cent). LS (39 per cent), MU (33 per cent), and PU (50 per cent), whereas [U-14C]-glutamine decarboxylation was increased by LS (53 per cent) and MU (55 per cent) and decreased by MS (17 per cent). The results presented indicate that the reduction in lymphocyte proliferation due to LS, LU and PU could well be a consequence of changes in glucose and glutamine metabolism.

Dietary fats alter the activity and expression of glucose-6-phosphate dehydrogenase in rat lymphoid cells and tissues.

The effect of diets enriched with fat containing different fatty acids on the activity and expression of the glucose-6-phosphate dehydrogenase (EC 1.1.1.49) of mesenteric lymph nodes lymphocytes and intraperitoneal macrophages was examined. Measurements of the enzyme were also performed using spleen, thymus and liver for comparison. The following fat rich diets containing a variety of fatty acids were used: 1-standard chow (CC); 2-medium chain saturated fatty acids (MS)-coconut fat-oil; 3-long chain saturated fatty acids (LS)-cocoa butter; 4-monounsaturated fatty acids (MU)-canola oil (n-9); 5-polyunsaturated fatty acids (PU)-soybean oil (n-6). Of the fat-rich diets tested, MS had the least effect. The G6PDH activity of lymphocytes was reduced by all the fat-rich diets; 16% for MS, 38% for LS, and 54% for MU. Similarly, the enzyme activity was reduced in macrophages; 35%, 86%, and 73%, for LS, MU, and PU, respectively. In contrast, the fat-rich diets elevated G6PDH activity in the lymphoid organs; by 42% in the spleen due to LS and by 131%, 35%, and 56% in the thymus due to LS, MU, and PU, respectively. Fat-rich diets decreased the activity of G6PDH in liver; 42%, 68%, and 39% for MS, MU, and PU, respectively. Some of the changes in G6PDH activity induced by the fat-rich diets occur through the mechanisms of mRNA abundance.

Structure and function of a new STAT-induced STAT inhibitor.

The signalling pathway that comprises JAK kinases and STAT proteins (for signal transducer and activator of transcription) is important for relaying signals from various cytokines outside the cell to the inside. The feedback mechanism responsible for switching off the cytokine signal has not been elucidated. We now report the cloning and characterization of an inhibitor of STAT activation which we name SSI-1 (for STAT-induced STAT inhibitor-1). We found that SSI-1 messenger RNA was induced by the cytokines interleukins 4 and 6 (IL-4, IL-6), leukaemia-inhibitory factor (LIF), and granulocyte colony-stimulating factor (G-CSF). Stimulation by IL-6 or LIF of murine myeloid leukaemia cells (M1 cells) induced SSI-1 mRNA expression which was blocked by transfection of a dominant-negative mutant of Stat3, indicating that the SSI-1 gene is a target of Stat3. Forced overexpression of SSI-1 complementary DNA interfered with IL-6- and LIF-mediated apoptosis and macrophage differentiation of M1 cells, as well as IL-6 induced tyrosine-phosphorylation of a receptor glycoprotein component, gp130, and of Stat3. When SSI-1 is overexpressed in COS7 cells, it can associate with the kinases Jak2 and Tyk2. These findings indicate that SSI-1 is responsible for negative-feedback regulation of the JAK-STAT pathway induced by cytokine stimulation.

Effects of electrochemotherapy on CaSki cells derived from a cervical squamous cell carcinoma.

Electrochemotherapy is a new approach to the treatment of tumors that takes advantage of the permeabilization of the cell membranes by electric pulses to facilitate the intracellular delivery of chemotherapeutic drugs into cells. Using the female genital squamous cell carcinoma cell line, CaSki, the antitumor effectiveness of electrochemotherapy was tested. In vitro studies showed that the cytotoxicity of some anticancer drugs, especially bleomycin, can be greatly enhanced by exposing cells to noncytotoxic electric pulses. This enhancement was significantly greater when the electric treatment was given after exposure to the drug than when applied preexposure. Treatment of nude mice bearing subcutaneous transplanted tumors with noneffective intraperitoneal doses of bleomycin followed by local delivery of electric pulses similar to those performed in vitro resulted in tumor reduction and complete disappearance after 12 days this electrochemotherapy. Histological changes in tumor tissue such as necrosis and degeneration were extensive even 6 hr after the electrochemotherapy. Thus the antitumor effects of bleomycin in mice can be considerably potentiated by local electric pulses, suggesting that electrochemotherapy with bleomycin may have promise for treatment of vulvar or ectocervical squamous cell carcinomas with early invasion.

Carnitine depletion during total parenteral nutrition despite oral L-carnitine supplementation.

Carnitine CAR) plays an important role in the beta-oxidation of fatty acids. Less attention. however, has been paid to CAR compared to other nutrients even in total parenteral nutrition (TPN). To examine CAR metabolism during TPN and the effect of simultaneous oral L-CAR supplementation on CAR levels, the blood CAR level was measured in a 3-year-old boy receiving long-term TPN because of short bowel syndrome. Both the total and acyl CAR in the serum were evaluated under various nutritional conditions including oral supplementation of L-CAR. Low CAR concentrations were observed especially when lipid containing TPN regimens were in place. Oral L-CAR supplementation was not sufficient to restore the low CAR levels in the present index patient even when the dose was increased to 120 mg/kg in accordance with the result of the L-CAR absorption test that revealed poor intestinal absorption of this nutrient. Moreover, a markedly low CAR level was measured during the onset of sepsis in the patient, and the blood CAR was depleted when lipid metabolism was activated by lipid loading or sepsis. To date, the late effects of CAR depletion on child growth have not been well examined. It is recommended that the blood CAR level be maintained at normal levels before any prominent manifestations of the deficiency have developed. The intravenous administration of CAR appears to be necessary to supply a sufficient amount of CAR for patients with severe malabsorption.

A new inositol 1,4,5-trisphosphate binding protein similar to phospholipase C-delta 1.

We have reported that two inositol 1,4,5-trisphosphate binding proteins, with molecular masses of 85 and 130 kDa, were purified from rat brain; the former protein was found to be the delta 1-isoenzyme of phospholipase C (PLC-delta 1) and the latter was an unidentified novel protein [Kanematsu, Takeya, Watanabe, Ozaki, Yoshida, Koga, Iwanaga and Hirata (1992) J. Biol. Chem. 267, 6518-6525]. Here we describe the isolation of the full-length cDNA for the 130 kDa Ins(1,4,5)P3 binding protein, which encodes 1096 amino acids. The predicted sequence of the 130 kDa protein had 38.2% homology to that of PLC-delta 1. Three known domains of PLC-delta 1 (pleckstrin homology and putative catalytic X and Y domains) were located at residues 110-222, 377-544 and 585-804 with 35.2%, 48.2% and 45.8% homologies respectively. However, the protein showed no PLC activity to phosphatidylinositol 4,5-bisphosphate and phosphatidylinositol. The 130 kDa protein expressed by transfection in COS-1 cells bound Ins(1,4,5)P3 in the same way as the molecule purified from brain. Thus the 130 kDa protein is a novel Ins(1,4,5)P3 binding protein homologous to PLC-delta 1, but with no catalytic activity. The functional significance of the 130 kDa protein is discussed.

A case of drug eruption caused by the crude drug Boi (Sinomenium stem/Sinomeni caulis et Rhizoma).

We report a case of drug eruption caused by the crude drug Boi. A 41-year-old female patient had been diagnosed with chronic rheumatoid arthritis in the department of internal medicine. After ingestion of a decoction of the crude drug Boi for the alleviation of arthralgia, a slight fever developed, which was followed by systemic edematous erythema with itching. HPLC showed that the main components of the crude drug Boi are sinomenine and magnoflorine. The results of patch tests were negative for all oral drugs that the patient had been taking. Oral ingestion tests showed that the patient showed positive reactions to the as-is Boi boiling-water decoction and 1/10-volume sinomenine. Based on this, the drug eruption was judged to be caused by sinomenine. It is considered the first time that the causative component of a drug eruption was confirmed by oral ingestion tests with components of a crude drug of Kampo medicine (Sino-Japanese traditional medicine).

Pheophorbide a, a potent endothelin receptor antagonist for both ETA and ETB subtypes.

Many crude drugs were screened for their capacity to inhibit the binding of endothelin-1 (ET-1) to ET receptors; several crude drugs showed significant binding inhibitory activity. Pheophorbide a (1), a potent non-peptide ET receptor antagonist, was isolated from Altemisiae capillaris Flos ("Inchinko" in Japanese), which has been utilized as a remedy for hepatitis in Oriental medicine. In receptor binding experiments, compound 1 inhibited ET-1 binding specifically to both the ETA receptor (ETAR) and ETB receptor (ETBR), with IC50 values of 8.0 x 10(-8) and 2.1 x 10(-7) M, respectively. Thus, compound 1 is an ET-1 binding inhibitor; however, it exhibited no affinity for the other receptors of angiotensin II and atrial natriuretic peptide. We also evaluated the inhibitory activity of porphyrin compounds, and found that some exhibited moderate activity.

Low prevalence of hepatitis C virus infection among hospital staff and acupuncturists in Kyushu, Japan.

During 1987 and 1988, samples of serum were collected from 1097 members of the staff of four prefectural hospitals in Miyazaki prefecture and from 183 acupuncturists in Fukuoka City, Japan. The staff included both surgical and non-surgical doctors, radiographers, physiotherapists, nurses, laboratory technicians and others. The samples were tested for the following hepatitis C virus (HCV) markers; antibodies to c100 (anti-c100) by means of enzyme-linked immunosorbent assay (ELISA) with supplementary recombinant immunoblot assay as well as antibodies to the GOR epitope (anti-GOR), also by means of ELISA. Anti-c100 was present in 1.7% of the doctors, radiographers and physiotherapists, in 1.3% of the nurses and in 2.2% of the acupuncturists. These prevalences were slightly higher than those in the controls but the differences were not statistically significant. Anti-c100 was not detected in any laboratory technician or other member of the hospital staff. For an accurate determination of the prevalence of HCV infection, the combined rate of anti-c100 and/or anti-GOR was also calculated. The combined prevalence of HCV infection was 4.3% in medical staff, 2.2% in nurses and 5.5% in acupuncturists. The prevalence of HCV infection among those with direct contact with patients was higher than that of the controls but without statistical significance. In the cohort we examined, the occupational risk of HCV infection was low.

[The influence of serum separators on biochemical values in experimental animals].

The influence of three serum separators, A, B and C, on biochemical values was examined. With A or B, changes in the values of ChE, LAP, UN, K, P, Cl and Ca in rat serum; ChE, UN, Cl and Ca in dog serum; and K and P in monkey serum took place over a period of 20 min after blood collection. Therefore, the biochemical values of whole blood were considered to be stable after 20 min. Thus, biochemical tests were conducted on serum from the three serum separators after allowing the blood to stand 20 min. The values obtained for each separator were not markedly different from those of the control. These results suggest that a serum separator is useful for separation of serum of experimental animals under the proper conditions.

Decreased hypothalamic and medullary GABA turnover in spontaneously hypertensive rats.

OBJECTIVE: The aim was to assess whether Gamma-aminobutyric acid (GABA) neurone activities in the central nervous system, especially in the hypothalamus and medulla oblangata, are altered in hypertension. METHODS: Central GABA content and turnover rate were measured in spontaneously hypertensive rats (SHR) and their normotensive Wistar Kyoto controls (WKY). GABA content was determined with high performance liquid chromatography, and in vivo GABA turnover rates were estimated by GABA accumulation after injection of amino-oxyacetic acid, a selective inhibitor of GABA degrading system. Two groups of nine week old male rats (32 SHR and 32 WKY) were used. RESULTS: GABA concentrations in cerebrospinal fluid were lower in SHR than in WKY. Since hypothalamus and medulla oblongata are the possible active sites of this system, basal GABA contents and in vivo GABA turnover rates were measured in hypothalamus and medulla oblongata. Basal GABA content in the medulla oblongata and hypothalamus was almost equal in SHR and WKY. On the other hand, GABA turnover rates were significantly lower in SHR than in WKY in both the hypothalamus and the medulla. CONCLUSIONS: Since it is known that GABA is an inhibitory neurotransmitter in the central nervous system and that it controls autonomic and cardiovascular activities, the findings suggest that the decreased hypothalamic and medullary GABAergic activities may permit sympathetic hyperactivity to contribute to the increase in blood pressure in SHR.