Effect of acute and chronic cholinesterase inhibition on biogenic amines in rat brain

The effects of five cholinesterase inhibitors on forebrain monoamine and their metabolite levels, and on forebrain and plasma cholinesterase (ChE) activity in rat were studied in acute and chronic conditions. Acute tetrahydroaminoacridine (THA) dosing caused lower brain (68%) and higher plasma (90%) ChE inhibition than the other drugs studied increased levels of brain dihydroxyphenylacetic acid (DOPAC) (236%), homovanillic acid (HVA) (197%) and 5-hydroxyindolaecetic acid (5-HIAA) (130%). Acute physostigmine (PHY) administration caused a 215% increase in brain DOPAC content. Despite high brain ChE inhibition induced by metrifonate (MTF), dichlorvos (DDVP) or naled no changes in brain noradrenaline (NA), dopamine (DA) or serotonin (5-HT) occurred due to treatment with the study drugs in the acute study. In the chronic 10-day study THA or PHY caused no substantial ChE inhibition in brain when measured 18 hours after the last dose, whereas MTF induced 74% ChE inhibition. Long-term treatment with THA or MTF caused no changes in monoamine levels, but PHY treatment resulted in slightly increased 5-HT values. These results suggest that MTF, DDVP and naled seem to act solely by cholinergic mechanisms. However, the central neuropharmacological mechanism of action of THA and PHY may involve changes in cholinergic as well as dopaminergic and serotoninergic systems.     

Studies on new,centrally active and reversible acetylcholinesterase inhibitors

We have synthesized the tertiary amines of pyridostigmine and neostigmine, 3-pyridinol dimethylcarbamate (norpyridostigmine) and 3-dimethylaminophenol dimethylcarbamate (norneostigmine) respectively, and we have tested their abilities to cross the blood-brain barrier and inhibit mouse brainAChE activity. The in vivo inhibition of AChE activity by norpyridostigmine reaches 72% at 10 minutes which is comparable to that seen with physostigmine (73% at 10 minutes). Inhibition by norneostigmine is less effective (50% at 10 minutes) and approaches that obtained with tetrahydroaminoacridine (57% at 10 minutes). These data show that both norpyridostigmine and norneostigmine cross the blood-brain barrier and that they are effective inhibitors of mouse brain AChE activity. These drugs could be useful in the treatment of memory, impairment associated with Alzheimer's disease, and other memory disorders.     

东莨菪碱镇痛作用的研究

腹腔(25或50mg/kg)或侧脑室注射东莨菪碱(1mg/kg),均使大鼠的痛阈升高42—80%,并持续32—36小时之久。当在恒温室内作昼夜连续观察时,对照组的大鼠在深夜20时至次日8时之前,痛刺激反应稍有增强,此时东莨菪碱组的镇痛作用也相应地减弱,而白天的镇痛作用较强,似有昼夜规律的表现。东莨菪碱的镇痛作用与脑内阿片能系统无关,DA 能暂时增强其镇痛作用,毒扁豆碱能拮抗其镇痛作用,所以,东莨菪碱的镇痛作用,主要是阻滞中枢 M-胆碱能系统的作用所致。     

Effect of oxotremorine,physostigmine, and scopolamine on brain acetylcholine synthesis: A study using HPLC

The synthesis rate of brain acetylcholine (ACh) was estimated in mice following i.v. administration of [³H]choline (Ch). The measurements were performed 1 min after the tracer injection, using the [³H]ACh/[³H]Ch specific radioactivity ratio as an index of ACh synthesis rate. Endogenous and labeled Ch and ACh were quantified using HPLC methodology. Oxotremorine and physostigmine (0.5 mg/kg, i.p.) increased the steady state concentration of brain ACh by +130% and 84%, respectively and of Ch by +60% (oxotremorine); they decreased ACh synthesis by 62 and 55%, respectively. By contrast, scopolamine (0.7 mg/kg, i.p.) decreased the cerebral content of Ch by –26% and of ACh by –23% without enhancing the synthesis of ACh. The results show the utility of HPLC methodology in the investigation of ACh turnover.     

Thermal resistance of the ciliary activity in the gills of the fresh water mussel Anodonta anatina

1. 1.|The thermal resistance of the activity of frontal cilia in the median gills of the fresh water mussel Anodonta anatina was studied.

2. 2.|The resistance acclimation appeared in 2 days in the gills of intact animals, but not in the isolated gills kept at 4, 14 and 24°C, for between 1 to 3 days.

3. 3.|Warm-acclimation increased the ACh sensitivity of the gills of intact mussels.

4. 4.|Isolation of the gills enhanced the thermal resistance.

5. 5.|ACh, choline and tetramethylammonium enhanced the thermal resistance in the isolated gills. whereas atropine and physostigmine diminished it.

6. 6.|It is concluded that in A. anatina the control if the thermal resistance is probably neural.

Does cholinesterase participate in the intercellular interactions in pollen-pistil system?

Hydrolysis of acetylthiocholine and butyrylthiocholine has been observed in aqueous extracts from petunia pollen and pistils. The reproductive organs of self-compatible clone showed a higher rate of choline ester hydrolysis than those of self-incompatible clone. The highest rate of acetylthiocholine hydrolysis blocked by the cholinesterase inhibitors (physostigmine and neostigmine) was characteristic for the pollen of self-compatible clone. The incomplete (25 - 40 %) inhibition of hydrolysis in pistil extracts of self-compatible clone suggests the presence of unspecific esterases. The eight-fold lower hydrolysis was observed in the pistils of self-incompatible clone as compared to the pistils of compatible clone; neostigmine completely blocked this low hydrolytic activity. The treatment of flower buds with physostigmine and neostigmine (10-5 - 10-3 M) decreased the seed production by 10 - 20 % in compatible clone. When the surfaces of pistil stigmae were treated with physostigmine and neostigmine (10-5 - 10-3 M) before pollination, the seed formation was inhibited by 95 % after both self- and cross-pollination. This revised version was published online in July 2006 with corrections to the Cover Date.     

Physostigmine production byStreptomyces griseofuscus NRRL 5324: Process development and scale-up studies

A reliable and scalable fermentation process was developed for production of the acetylcholine esterase inhibitor physostigmine employingStreptomyces griseofuscus NRRL 5324. Initial fermentation in small-scale bioreactors reached physostigmine levels of approximately 60 mg L^–1 after 139 h. Optimization of both process operating parameters and production medium composition rapidly yielded a seven-fold increase in physostigmine titer. The scaled up process routinely produced physostigmine titers of approximately 400 mg L^–1 during a fermentation cycle of 180 h, and supported the rapid production of large amounts of physostigmine. A physostigmine production of 500 mg L^–1 representing an eight-fold improvement over the original performance, was achieved using a glucose/ammonium fed-batch process.     

兔右房超速驱动后阻抑机制的初步探讨

本文用微电极技术,在20份离体兔心窦房结标本上观察了不同驱动频率和驱动时程对窦房结超速驱动后阻抑的影响。结果表明,在一定范围内右房超速驱动后阻抑及驱动后的负性变速效应,随驱动频率和驱动时程增加而加强。高频驱动期间和驱动停止后,优势起搏细胞跨膜电位幅度减小。驱动停止后第一个窦性动作电位4相自动去极化斜率和0相上升速率均降低,与驱动前相比有非常显著的差异(P<0.01)。毒扁豆碱可加强驱动后阻抑及驱动后的负性变速效应,阿托品不能完全阻断驱动后阻抑。我们初步认为:右房超速驱动后阻抑效应可能是内源性乙酰胆碱的释放和[Na~ ]_i 及[Ca~2 ]_i 的增加等多种因素作用的结果。     

Metrifonate induces cholinesterase inhibition exclusively via slow release of dichlorvos

Metrifonate, a long-acting cholinesterase (ChE) inhibitor with very low toxicity in warm-blooded animals, inhibits rat brain and serum cholinesterase (ChE) in vitro through its hydrolytic degradation product, dichlorvos. This conclusion is based on the finding that metrifonate-induced ChE inhibition showed the same pH dependence as its reported dehydrochlorination to dichlorvos. The ChE inhibition induced by dichlorvos was not pH dependent. It was mediated by a competitive drug interaction with the catalytic site of the enzyme, which led to irreversible inhibition within several minutes of incubation. After this time, addition of further substrate to the inhibited enzyme was not able to promote drug dissociation and hence enzyme reactivation. Similar characteristics of inhibition, i.e. interaction with the substrate binding site and time-dependent switch to non-competitive inhibition were observed with the reference compound, physostigmine. However, the physostigmine-induced inhibition of ChE could be readily reversed by further substrate addition. Another reference compound, tetrahydroaminoacridine (THA), also induced a reversible inhibition of rat brain and serum cholinesterase, but with a mechanism of action different from that of both dichlorvos and physostigmine in that enzyme inhibition occurred rapidly upon drug addition at an allosteric site on the enzyme surface. It is suggested that the unique slow release plus the slow inhibition of ChE by dichlorvos is responsible for the lower toxicity of metrifonate compared to that of directly acting ChE inhibitors.