Nicotine at less than or equal to 33 microM enhanced the single twitch response to indirect stimulation but potentiated the blocking effect of tubocurarine. Failure of tetanic contraction (tetanic fade) occurred on stimulation at 100 Hz. At 76 microM, nicotine induced a first phase rapid (10 min) inhibition of twitch response followed later (60-90 min) by a second phase complete block. Neostigmine partially restored the response at either phase of block whereas diaminopyridine completely antagonized the blockade. The end-plate was depolarized maximally by only 10-15 mV within 30 min with 43 microM nicotine. The depolarization was maintained but was antagonized by tubocurarine. The twitch response induced by direct stimulation was unchanged indicating no depolarization block ensued. The amplitudes of both EPP (0.7 Hz) and MEPP were markedly depressed in parallel indicating a curare-like postsynaptic inhibition without an effect on the release of transmitter. It is concluded that nicotine blocks the neuromuscular transmission by a dual mechanism by its partial agonist action. At higher frequencies of transmission, nicotine (greater than or equal to 22 microM) also produced a remarkable run-down of EPP just like other receptor antagonists suggesting that the nerve terminal acetylcholine receptors are not particularly sensitive to nicotine as those on the autonomic ganglia.

4-Aminopyridine↗

PubMed1987

Cytologic presentation and its correlation with clinical stage in papillary carcinoma of the thyroid.

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Biopsy, Needle↗

PubMed1986

Biosynthesis of monoterpenes. Enantioselectivity in the enzymatic cyclization of (+)- and (-)-linalyl pyrophosphate to (+)- and (-)-bornyl pyrophosphate.

Enzymes from Salvia officinalis and Tanacetum vulgare leaf epidermis catalyze the conversion of the acyclic precursor geranyl pyrophosphate to the cyclic monoterpenes (+)- and (-)-bornyl pyrophosphate, respectively. The antipodal cyclizations are considered to proceed by the initial isomerization of the substrate to the respective bound tertiary allylic intermediates (-)-(3R)- and (+)-(3S)-linalyl pyrophosphate. [(3R)-8,9-14C,(3RS)-1E-3H] Linalyl pyrophosphate (3H:14C = 5.22) was tested as a substrate with the cyclases from both sources to determine the configuration of the cyclizing intermediate. This substrate yielded (-)-bornyl pyrophosphate with 3H:14C ratio greater than 31, indicating specific utilization of (+)-(3S)-linalyl pyrophosphate as predicted. With the (+)-bornyl pyrophosphate cyclase, the 3H:14C ratio of the product was about 4.16, indicating a preference for the (-)-(3R)-enantiomer, but the ability also to utilize (+)-(3S)-linalyl pyrophosphate. (3R)- and (3S)-[1Z-3H]Linalyl pyrophosphate were separately compared to the achiral precursors [1-3H] geranyl pyrophosphate and [1-3H]neryl pyrophosphate (cis-isomer) as substrates for the cyclizations. All functional precursors afforded optically pure (-)-(1S,4S)-bornyl pyrophosphate with the T. vulgare-derived cyclase (as determined by chromatographic separation of diastereomeric ketals of the derived ketone camphor), and (+)-(3S)-linalyl pyrophosphate was the preferred substrate. With the (+)-bornyl pyrophosphate cyclase from S. officinalis, geranyl, neryl, and (-)-(3R)-linalyl pyrophosphates gave the expected (+)-(1R,4R)-stereoisomer as the sole product, and (-)-(3R)-linalyl pyrophosphate was the preferred substrate. However, (3S)-linalyl pyrophosphate yielded (-)-(1S,4S)-bornyl pyrophosphate, albeit at lower rates, indicating the ability of this enzyme to catalyze the anomalous enantiomeric cyclization.

Acyclic Monoterpenes↗

PubMed1986

A regenerating release of acetylcholine from mouse motor nerve terminals treated with anticholinesterase agents.

It was found by intracellular recording with glass microelectrodes that train stimulation (50-200 Hz) of the phrenic nerve of intact or cut mouse diaphragm induced an accumulative depolarization of the endplate and triggered after a few pulses an 'all-or-none' regenerative depolarization lasting for 300-900 ms when acetylcholinesterase was inhibited by neostigmine or diisopropylfluorophosphate. This depolarization was associated with a noise of the membrane potential and a failure of the end plate potential. Low Ca2+ prolonged whereas high Ca2+ shortened the duration of regenerative depolarization which needed no further stimulation once triggered. d-Tubocurarine abolished the depolarization while restoring the end plate potential. A regenerative release of acetylcholine due to an activation of presynaptic cholinoceptors is speculated.

Acetylcholine↗

PubMed1986

Cycloheximide sensitivity in regulation of acyl coenzyme A:cholesterol acyltransferase activity in Chinese hamster ovary cells. 1. Effect of exogenous sterols.

Chinese hamster ovary cells grown in medium containing low-density lipoprotein (LDL) express high acyl coenzyme A:cholesterol acyltransferase (ACAT) activity as measured by an [3H]oleate pulse. Removal of LDL from the medium causes rapid inactivation of ACAT activity; the t1/2 for the initial inactivation rate is 0.8 h. Preincubation with protein synthesis inhibitors (cycloheximide or emetine) for 2 h or longer lengthens the t1/2 for the initial inactivation rate to approximately 2.1 h. When LDL is removed for more than 10 h, the cells contain only 3% of the original ACAT activity. Cycloheximide under this condition causes an 8-fold increase in ACAT activity; the increase approaches a maximum in 6-8 h. The extent of ACAT activation by cycloheximide inversely depends on exogenous sterol present in the medium; LDL diminishes the activation, while cationized LDL or 25-hydroxycholesterol completely abolishes the activation. Adding LDL back to the sterol-free medium causes a 40-70-fold increase in ACAT activity; however, the activation of LDL is not further augmented if the cells are pretreated with cycloheximide. The above observations are qualitatively confirmed by ACAT assays in vitro with cell homogenates. LDL or cycloheximide has no effect on the rates of 3H-labeled triglyceride and 3H-labeled polar lipid synthesis. Efflux of prelabeled cholesterol from cells is cycloheximide-insensitive. Rates of degradation of [3H]-leucine-pulse-labeled total protein in cells grown with or without LDL are identical. The above results imply the existence of at least one specific short-lived factor that directly or indirectly inhibits ACAT activity.(ABSTRACT TRUNCATED AT 250 WORDS)

Animals↗

PubMed1986

Cycloheximide sensitivity in regulation of acyl coenzyme A:cholesterol acyltransferase activity in Chinese hamster ovary cells. 2. Effect of sterol endogenously synthesized.

We reported in another paper [Chang, C. C. Y., Doolittle, G. M., & Chang, T. Y. (1986) Biochemistry (preceding paper in this issue)] that in Chinese hamster ovary (CHO) cells activation of acyl coenzyme A:cholesterol acyltransferase (ACAT) activity by treating cells with cycloheximide was abolished by providing exogenous sterol in the medium. We now report that providing 20 mM DL-mevalonate to cells grown in sterol-free medium increases the ACAT activity by approximately 6-fold and diminishes the cycloheximide activation effect. The mevalonate supplement has no significant effect on the rate of triglyceride or polar lipid synthesis, [3H]cholesterol efflux, or bulk protein degradation in cells. The activation of ACAT by mevalonate is prevented by adding a specific squalene oxide cyclase inhibitor to cells, indicating the requirement for endogenous sterol synthesis to mediate the mevalonate effect. In sterol-free medium, if sterol synthesis is blocked by specific enzyme inhibitors or through mutation, the ACAT activation by cycloheximide is again abolished. These results support the hypothesis that there may exist a short-lived factor(s) serving directly or indirectly as an endogenous ACAT inhibitor(s), the inhibitory action of this (these) factor(s) is (are) abolished, and its (their) turnover rate(s) is (are) increased by either exogenous sterol or by sterol endogenously synthesized. Conversely, removing exogenous sterol coupled with blocking endogenous sterol synthesis decreases the turnover rate(s) of the inhibitor(s), rendering its (their) action insensitive to intracellular degradation over the time period studied.

Animals↗

PubMed1986

Inhibition of gap junctional intercellular communication in human teratocarcinoma cells by organochlorine pesticides.

Inhibition of intercellular communication, as measured by metabolic cooperation between 6-thioguanine-sensitive and 6-thioguanine-resistant Chinese hamster V79 cells, has been previously shown to be correlated with a large variety of known tumor promoters, including some of the organochlorine pesticides. Since further evidence concerning the effects of those known or suspected animal tumor promoters on human cells is needed, three organochlorine pesticides, dieldrin, aldrin, and 1,1,1-trichloro-2,2-bis(p-chlorophenyl)ethane (DDT), were tested for their ability to inhibit metabolic cooperation between 6-thioguanine-sensitive (6TGs, HTP3-4) and 6-thioguanine-resistant (6TGr, HTXTG-1) human teratocarcinoma cells. Similar to the effect of the known mouse skin tumor promoter 12-tetradecanoyl phorbol-13-acetate (TPA), all three pesticides inhibited gap junctional intercellular communication within a noncytotoxic dose range. The dose-response curves of these chemicals were similar to those of other known tumor promoters on Chinese hamster V79 cells. In addition, the transfer of [3H]uridine between teratocarcinoma cells in contact was reduced after pesticide treatment.

Aldrin↗

PubMed1986

Role of intercellular communication in modifying the consequences of mutations in somatic cells.

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Animals↗

PubMed1986

Pharmacological identification of saxitoxin-like toxins in the cultured purple clam, Hiatula diphos.

An outbreak of poisoning from ingestion of purple clam (Hiatula diphos), obtained from a brackish fish-culture pond, occurred in southern Taiwan in early January 1986. The extract blocked the twitch responses and generation of the action potential of the mouse diaphragm without an effect on the resting membrane potential or the miniature endplate potential. The depolarizations and spontaneous firings of the diaphragm induced by veratridine and crotamine were effectively antagonized by the toxic clam extract, indicating the presence of saxitoxin-like substances, which may have amounted to 0.013% tetrodotoxin equivalent.

Animals↗

PubMed1986

Formulation of a potential antipregnancy vaccine based on the beta-subunit of human chorionic gonadotropin (beta-hCG). IV. The role of adjuvants in booster injections.

Experiments were conducted in rabbits to determine the effect of adjuvant use on the antibody response following booster injections. The antigen used was in all cases the beta-subunit of human chorionic gonadotropin linked to tetanus toxoid (beta-hCG-TT). Adjuvants used were Al(OH)3, MDP analogs, and a streptococcus preparation, OK432. Primary vaccinations included Al(OH)3 adjuvant with or without supplementary adjuvants. In general, the greater the antibody response following primary vaccination, the greater the response following booster vaccination whether or not adjuvant was used in the booster. No increment in antibody titers was found by reason of including MDP analogs in booster vaccinations. OK432, in contrast, gave increased responses in booster injections which were in several cases statistically significant. The value of including Al(OH)3 in booster injections is not clear from the experimental data. In no case was the increment due to its inclusion large.

Adjuvants, Immunologic↗

PubMed1986

Mechanisms of the inhibition by neostigmine of tetanic contraction in the mouse diaphragm.

Neostigmine (0.5-2 microM) caused fade of tetanic contractions (Wedensky inhibition) evoked by repetitive nerve stimulation. The mechanism underlying this action was studied in intact and cut isolated phrenic nerve-diaphragm preparations of mice. The fade was brought about by failure to elicit muscle action potentials. During fade, the muscle was unable to conduct directly evoked action potentials across the central endplate zone. Recovery of excitability occurred in 5 s with continued stimulation. In the presence of neostigmine, the resting membrane potential at endplate areas during repetitive stimulation decreased from -80 mV to less than -50 mV within the first 10 pulses at 75-200 Hz and thereafter recovered gradually to about -60 mV in the following 5 s during continuous stimulation. The quantal content of endplate potentials evoked by single stimulation was not reduced by neostigmine whereas that evoked by high frequency stimuli (75 Hz) was reduced to about 1/3 in 10 pulses. It is concluded that the fade of tetanic contraction caused by inhibition of acetylcholinesterase is induced by the inactivation of sodium channels in the area surrounding the endplates and that the sustained fade is due to a decrease of transmitter release. Both effects are the result of acetylcholine accumulation.

Action Potentials↗

PubMed1986

A comparison of the antagonisms by neostigmine and diaminopyridine against the neuromuscular block caused by cobrotoxin and (+)-tubocurarine.

Cobrotoxin was about 11-fold more potent than (+)-tubocurarine on a weight basis in blocking neuromuscular transmission in mouse isolated phrenic nerve-diaphragm preparations. Neostigmine and diaminopyridine increased the concentrations of cobrotoxin for 70% inhibition of indirect contraction by 290 and 320%, and increased those of (+)-tubocurarine by 180 and 230%, respectively. More than additive increases were obtained when neostigmine and diaminopyridine were used simultaneously. Cobrotoxin, however, was only 6-fold more toxic than (+)-tubocurarine after intraperitoneal injection in mice. The lethal dose of (+)-tubocurarine was increased by 80% when both antidotes were used together, but only by 15-20% when used alone. In contrast, the lethality of cobrotoxin was not decreased by these drugs. Unexpectedly, the time to death after treatment with cobrotoxin was shortened when mice were pretreated with these antidotes.

4-Aminopyridine↗