Evidence for the involvement of protein kinase C isoforms in alpha-1 adrenergic activation of phospholipase A2 in FRTL-5 thyroid cells

BACKGROUND: FRTL-5 thyroid cells are a cell line extensively used for the investigation of thyroid functions. Activation of alpha-1 adrenergic receptors stimulates both arachidonic acid (AA) release and cytosolic Ca2+ increase in this cell line. Cytosolic Ca2+ and arachidonic acid are known to be important second messengers regulating a variety of thyroid functions. The generation of these messengers is regulated primarily by two different types of phospholipases, phospholipase C (PLC) and phospholipase A2 (PLA2). METHODS: Norepinephrine (NE, 10 mumol/L) was used as an alpha-1 adrenergic activator, and cytosolic-free Ca2+ concentration ([Ca2+]i) was determined using the fluorescent dye indo-1. Arachidonic acid release was measured as an indicator of PLA2 activation, and protein kinase C (PKC) activity determination and isoforms identification were performed using commercial kits. RESULTS: Norepinephrine increased [Ca2+]i and AA release. Prevention of NE-induced cytosolic Ca2+ influx, either by removal of extracellular Ca2+ or by use of Ca2+ channel blockers, NiCl2 or CoCl2, inhibited AA generation entirely. Inhibition of NE-induced increase in [Ca2+]i by the Ca2+ chelator, 1,2-bis(2-aminophenoxy)ethane-N,N,N',N'-tetraacetic acid (BAPTA), also significantly suppressed NE-induced AA release. Inhibition of PKC activity by PKC inhibitors (H-7 or staurosporine) or downregulation induced by prolonged treatment with phorbol 12-myristate 13-acetate (PMA) or thyleametoxin (TX) significantly blocked the NE-induced AA release, which indicates PKC is involved in mediating NE-induced AA release. Protein kinase C activity measurement indicated that NE induced an activation of PKC in 5 minutes. To further characterize the role of PKC or Ca2+ in regulation of AA release, we identified PKC isoforms by immunoblotting with specific antibodies against 8 different Protein kinase C isoforms. PKC-alpha, -beta I, -beta II, -gamma, delta, -epsilon, -zeta, and -eta isoforms were identified. Norepinephrine induced translocation of PKC-alpha, -beta I, -beta II, -gamma, -delta, and -epsilon isoforms but not -zeta and -eta from cytosol to membrane. Chelation of intracellular Ca2+, prevention of Ca2+ influx, or prolonged treatment with thymeleatoxin (TX) completely blocked the NE-induced translocation of PKC-alpha. CONCLUSIONS: These results, taken together with data obtained from AA experiments, suggest that PKC plays a critical role in alpha-1 adrenergic receptor mediated PLA2 activation and subsequent AA release. Extracellular Ca2+ influx is a prerequisite for both PKC-alpha translocation and AA release. Whether Ca2+ acts directly upon the PLA2, or via PKC-alpha, to regulate AA generation is an intriguing question that remains to be clarified.     

A simple and inexpensive method for the identification of Staphylococcus epidermidis and Staphylococcus hominis

Eight hundred and ninety-two strains of Staphylococcus species were identified by means of desferrioxamine susceptibility and fermentation results of three carbohydrates, with the API Staph system (bioMérieux, France) as reference method. No identification could be obtained for 34 strains with API Staph. Of the remaining 858 strains, identical identification was obtained with 842 (98.1%). All 707 strains identified as Staphylococcus epidermidis or Staphylococcus hominis by the API Staph system were found to be desferrioxamine susceptible, and all but 5 (3.3%) of 151 strains identified as other staphylococcal species were found to be resistant, yielding an identification correlation of 99.4% for desferrioxamine. The five additional strains which were susceptible to desferrioxamine were identified as Staphylococcus capitis (2 strains), Staphylococcus lugdunensis (2 strains), and Staphylococcus warneri (1 strain) by API Staph, and as Staphylococcus epidermidis (1 strain), Staphylococcus hominis (3 strains), and one other staphylococcal species by the experimental system.     

Improved GaAs-based quantum cascade laser (/spl lambda//spl sim/11 /spl mu/m) using high-reflectivity metal facet coating

By applying a high-reflectivity metal coating to the rear facet of a GaAs-based quantum cascade laser operating at /spl lambda//spl sim/11.5 /spl mu/m, the threshold current has been reduced by 11% at 260 K and pulsed operation of the epilayer-up mounted device was extended from 283 to 294 K.     

Distal dorsal forearm flap

Reinstatement and spontaneous recovery of previously extinguished nicotine-taking behavior were examined in rats. Male subjects were trained to self-administer nicotine (30 microg/kg per infusion, IV; one 60-min session per day for 3 weeks). Extinction sessions were then given for 5-10 days during which saline was substituted for nicotine. Subsequently, in the first set of tests for nicotine seeking, the reinstatement of lever presses that previously delivered nicotine was examined after priming injections of saline and nicotine (75, 150 and 300 microg/kg, SC; and 30 and 60 microg/kg, IV). In the second set of tests for nicotine-seeking, rats were tested after an additional 21-day drug-free period during which they were not exposed to the self-administration chambers (a test for the spontaneous recovery of drug seeking), and after priming injections of nicotine (150 and 300 microg/kg, SC). Reinstatement of extinguished food-reinforced behavior after exposure to nicotine was also determined. Priming injections of nicotine reinstated nicotine seeking regardless of the route of administration. In addition, previously extinguished nicotine seeking recovered spontaneously after a 21-day period during which rats were not exposed to the drug-taking environment. Nicotine also reinstated extinguished food-reinforced behavior in rats with a history of nicotine self-administration, but not in drug-naive rats. The present results extend previous work with opioid and stimulant drugs on reinstatement of drug seeking by the self-administered drug. It also appears that, as with other positive reinforcers, the mere passage of time is a sufficient condition for the spontaneous recovery of extinguished nicotine seeking.     

Experimental investigation of corium quenching at elevated pressure

This paper describes the results of experiments designed to quantify the cooling rate of corium by an overlying water pool. The experiments are intended to provide fundamental information on the ability of water to ingress into cracks and fissures that form in the debris during quench, thereby augmenting the otherwise conduction-limited heat transfer process. This information is being used to assess the effectiveness of a water pool in thermally stabilizing a molten-core/concrete interaction and cooling of ex-vessel core debris. The experiments involved corium inventories of 75 kg with a melt depth of 15 cm and diameter of 30 cm. The corium was composed of UO₂/ZrO₂/concrete to simulate mixtures of molten reactor core components and either siliceous or limestone/common sand (LCS) concrete. Initial melt temperatures were of the order of 2100 °C. The heat transfer rate from the corium was determined through measurements of the vapor production rate from the water pool. The melt was quenched at atmospheric pressure for the first two tests and at 4 bar for the two subsequent tests. Preliminary data analysis indicates that the overall heat transfer rate exceeded the conduction-limited rate for the three melts containing 8 wt.% concrete, but not for the fourth, which had 23 wt.% concrete. Also, the quench rate of the 8 wt.% concrete melts did not vary appreciably with pressure.     

Recurrent hyperparathyroidism due to parathyroid autografts: incidence, presentation, and management

Recurrent hyperparathyroidism (HPT) occurs in a small percentage of patients undergoing parathyroidectomy for primary HPT and is usually due to inadequate excision of hyperfunctioning parathyroid tissue in the neck, a missed ectopic and hyperplastic parathyroid, or, less commonly, parathyroid carcinoma and parathyroid autografts. In order to determine the incidence, clinical characteristics, and outcome of patients with recurrent HPT due to parathyroid autografts, we reviewed our experience with 604 consecutive patients operated on for primary HPT between 1965 and 1989. One hundred of these patients received parathyroid autografts consisting of portions of one or more parathyroid glands. Three patients with autografts, placed in the sternocleidomastoid muscle, developed recurrent HPT due to their autografts for an incidence of 3 per cent. Recurrent disease was diagnosed between 62 and 113 months with an average of 89 months. The autotransplants in all three of these patients were from hyperplastic or adenomatous parathyroid tissue. Two patients had a history of neck irradiation. Preoperative thallium scans accurately localized the hyperfunctioning parathyroid tissue in all three patients. At operation, the hyperfunctioning autografts had grown into a discrete mass with a single vascular pedicle and were resected. Histologic examination disclosed either hyperplastic or adenomatous tissue, and corresponded to the histology and location of the original tissue transplanted in each case. Follow-up ranges from 12 to 67 months, with an average of 48 months. All patients remain cured and none require oral calcium supplementation. We conclude that graft-dependent recurrent HPT is due to the autotransplantation of hyperplastic or adenomatous parathyroid tissue and that thallium scanning is instrumental for diagnosis and localization.(ABSTRACT TRUNCATED AT 250 WORDS)