Phosphorylation and activation of tyrosine hydroxylase in PC18 cells: a cell line derived from rat pheochromocytoma PC12 cells.

Treatment of rat pheochromocytoma PC18 cells (a variant subclone of PC12 cells) with forskolin produced increased activity and phosphorylation of tyrosine hydroxylase. In contrast, treatment of the PC18 cells with 56 mM K+, A23187, phorbol-12-myristate-13-acetate (PMA) or phorbol-12,13-dibutyrate (PDB) did not affect the activity and only slightly increased the phosphorylation of tyrosine hydroxylase. None of the treatments except forskolin increased cyclic AMP levels in PC18 cells. Furthermore, 45Ca2+ uptake into PC18 cells was not affected by 56 mM K+, PDB or forskolin; however, A23187 increased 45Ca2+ uptake 4-fold over basal uptake. Nevertheless, no activation and little increase in phosphorylation of tyrosine hydroxylase was observed in PC18 cells treated with A23187. When tyrosine hydroxylase levels in PC18 cells were elevated by treatment with dexamethasone, activation of tyrosine hydroxylase by 56 mM K+, PDB or A23187 was still not observed. Both purified Ca2+/calmodulin-dependent protein kinase and cyclic AMP-dependent protein kinase catalyzed the phosphorylation of tyrosine hydroxylase purified from PC18 cells in vitro. Furthermore, crude cell extracts from PC12 cells and PC18 cells possessed Ca2+/calmodulin-dependent protein kinase activity that catalyzed the phosphorylation of purified tyrosine hydroxylase. These results suggest that tyrosine hydroxylase activity in PC18 cells is regulated by a cyclic AMP-dependent mechanism. However, due to a number of abnormalities the Ca(2+)-dependent mechanisms do not result in the activation of tyrosine hydroxylase and only slightly increase the phosphorylation of the enzyme in PC18 cells.     

Association of low caregiver health literacy with reported use of nonstandardized dosing instruments and lack of knowledge of weight-based dosing.

OBJECTIVE: Caregivers of young children frequently measure doses of liquid medications incorrectly. Use of nonstandardized dosing instruments and lack of knowledge that dosing is weight-based contribute to dosing errors. We sought to assess whether low caregiver health literacy was associated with these outcomes. METHODS: This was a cross-sectional analysis of caregivers presenting to an urban pediatric emergency room. Dependent variables were caregiver reported use of nonstandardized dosing tools and knowledge of weight-based dosing. The independent variable was caregiver health literacy (Test of Functional Health Literacy in Adults [TOFHLA]). RESULTS: Two hundred ninety-two caregivers were assessed: 23.3% reported use of nonstandardized liquid dosing instruments, and 67.8% were unaware of weight-based dosing. Caregivers who were unaware of weight-based dosing were more likely to use nonstandardized dosing tools (28.3% vs 12.8%; P = .003). In unadjusted analyses, overall health literacy, reading comprehension, and numeracy were all associated with both dependent variables. In analyses adjusting for child age, health care experiences, and caregiver acculturation and education, inadequate/marginal overall health literacy was associated with lack of knowledge of weight-based dosing (adjusted odds ratio [AOR] 2.3; P = .03), whereas lower reading comprehension was associated with both lack of knowledge (AOR 2.0; P = .03) and reported use of nonstandardized instrument (AOR 2.4; P = .007). CONCLUSIONS: Low health literacy, in particular reading comprehension, was associated with reported use of nonstandardized dosing instruments and lack of knowledge regarding weight-based dosing. Both caregiver health literacy and sociodemographic factors should be considered in the design of interventions to prevent medication administration errors.     

Extracorporeal surgery for renal tumour. A case report

A case of adenocarcinoma of a single functioning kidney is presented. The tumour was removed by extracorporeal bench surgery and the remnant successfully autotransplanted.     

Activation of a Ca2+-dependent K+ current by the oncogenic receptor protein tyrosine kinase v-Fms in mouse fibroblasts

We investigated the effects of the receptor-coupled protein tyrosine kinase (RTK) v-Fms on the membrane current properties of NIH3T3 mouse fibroblasts. We found that v-Fms, the oncogenic variant of the macrophage colony-stimulating factor receptor c-Fms, activates a K⁺ current that is absent in control cells. The activation of the K⁺ current was Ca²⁺-dependent, voltage-independent, and was completely blocked by the K⁺ channel blockers charybdotoxin, margatoxin and iberiotoxin with IC₅₀ values of 3nM, 18 nM and 76nM, respectively. To identify signalling components that mediate the activation of this K⁺ current, NIH3T3 cells that express different mutants of the wildtype v-Fms receptor were examined. Mutation of the binding site for the Ras-GTPase-activating protein led to a complete abolishment of the K⁺ current. A reduction of 76% and 63%, respectively, was observed upon mutation of either of the two binding sites for the growth factor receptor binding protein 2. Mutation of the ATP binding lobe, which disrupts the protein tyrosine kinase activity of v-Fms, led to a 55% reduction of the K⁺ current. Treatment of wild-type v-Fms cells with Clostridium sordellii lethal toxin or a farnesyl protein transferase inhibitor, both known to inhibit the biological function of Ras, reduced the K⁺ current amplitude to 17% and 6% of the control value, respectively. This is the first report showing that an oncogenic RTK can modulate K⁺ channel activity. Our results indicate that this effect is dependent on the binding of certain Ras-regulating proteins to the v-Fms receptor and is not abolished by disruption of its intrinsic protein tyrosine kinase activity. Furthermore, our data suggest that Ras plays a key role for K⁺ channel activation by the oncogenic RTK v-Fms. Received: 19 November 1997 / Accepted: 21 January 1998     

Distinct sites of action of clostridial neurotoxins revealed by double-poisoning of mouse motor nerve terminals

(1) We investigated the effects of single- and double-poisoning with tetanus toxin (TeTx), botulinum neurotoxin type A (BoTx A) and botulinum neurotoxin type B (BoTx B) on spontaneous and nerve-evoked quantal transmitter release at motor endplates of the triangularis sterni preparation of the mouse. (2) Inhibitory effects of TeTx and BoTx B on spontaneous and nerve-evoked transmitter release were very similar, except that the action of BoTx B required 500-fold lower concentrations and was less dependent on temperature. BoTx A caused stronger inhibition of quantal release than TeTx or BoTx B, but was comparatively much easier counteracted by 4-aminopyridine (4-AP). (3) In contrast to BoTx A, with TeTx or BoTx B the increase of transmitter release following onset of 50 Hz nerve stimulation was delayed for a few seconds and synaptic latencies of quanta showed large variations. This release pattern was also evident in all double-poisoning experiments, regardless of intoxication sequence. (4) Inhibition of evoked release was found to be slightly stronger with TeTx than with BoTx B, so the amount of nerve-evoked quanta released after double-poisoning with any sequence of these toxins always approached that of TeTx. In no case supraadditive actions were observed. (5) A strong reduction of evoked quanta was observed when BoTx A was applied in addition to either of the two other toxins. With reversed poisoning sequences (BoTx A-TeTx or BoTx A-BoTx B) the resulting values remained at the extremely low level of BoTx A. (6) In the presence of 4-AP double-poisoning with any combination between BoTx A and TeTx or BoTx B (regardless of intoxication sequence) revealed supra-additive effects, since the number of quanta released was considerably lower than that obtained with any of the toxins alone (in the presence of 4-AP). (7) Our results indicate that tetanus toxin and botulinum toxin type B have a common site of action which is different and independent from that of botulinum toxin type A.This is part of the thesis of M. G. to be presented to the Fachbereich Humanmedizin, Justus-Liebig-Universität Gießen