The effect of lanthanum ions on acetylcholine in frog muscle.

3. Spontaneous release of ACh varied in normal innervated muscles between 40 and 100 fmol/min. In the presence of 25 mM-KCl the rate of release increased about fourfold. In BoTx poisoned muscles spontaneous release was reduced by up to 60 % of control and high potassium failed to accelerate the release at 2 d after poisoning and caused only a small increase at 8 d. Denervated muscles released ACh at a rate which was less than 20 % of control and it was not accelerated by high potassium.4. The results show that more than 90 % oftotal ACh in the innervated EDL muscle is present in the nerve and its terminals. The remaining ACh is apparently formed and stored in the muscle tissue. BoTx caused a larger reduction in ACh release than can be accounted for by assuming a selective blockade of quantal release of transmitter. It suggests that BoTx has an inhibitory effect also on non-quantal ACh release.

Section: Discussionmentioning

confidence: 99%

“…According to estimates by Elmqvist & Quastel (1965) the average size of the presynaptic transmitter store in a single rat phrenic nerve terminal corresponds to 270,000 quanta. Assuming 70-80% of the total ACh in synaptic vessels (Heuser & Lennon, 1973;Miledi, Molenaar & Polak, 1980) one gets about 17,000 molecules of ACh per vesicle.…”

Section: Discussionmentioning

confidence: 99%

Acetylcholine content and release in denervated or botulinum poisoned rat skeletal muscle

Polak

Sellin

Thesleff

1981

“…Binding of 1251-aBTX was determined (Ito, Miledi, Vincent & Newsom-Davis, 1978) (Miledi, Molenaar & Polak, 1980) and measured using high-performance liquid chromatography (HPLC) with a method in which ACh was converted enzymically to H202 (Israel & Lesbats, 1981). The H202 was measured with a platinum electrode (Damsma, Lammerts van Bueren, Westerink & Horn, 1985).…”

Section: Biochemical Experimentsmentioning

confidence: 99%

Adaptation of quantal content to decreased postsynaptic sensitivity at single endplates in alpha‐bungarotoxin‐treated rats.

Plomp

Kempen

Molenaar

1992

Self Cite

173

149

SUMMARY1. Rats were injected once every 48 h with a-bungarotoxin (aBTX) for periods up to 6 weeks. Injections caused weakness of facial muscles which lasted about 8 h. Hemidiaphragms were dissected for biochemical and electrophysiological measurements.2. In muscles from animals treated for 2-3 weeks with toxin, the binding of 125-aBTX was reduced to 58 %, and the ACh content to 81 % of control values. Choline acetyltransferase activity was unchanged. ACh release evoked by 3 Hz nerve stimulation was increased to 175 % of control values.3. The use of ,u-conotoxin, which specifically blocks muscle action potentials, enabled the recording of full-sized endplate potentials (EPPs) and miniature endplate potentials (MEPPs) at normal muscle membrane potentials ( -70 to -80 mV). The amplitude of MEPPs was decreased to 57 % in muscles from animals treated for 3 weeks with aBTX. The mean of the quantal contents, calculated from the ratio of the corrected EPPs and the MEPPs, was increased to 154 %.4. Within individual muscles of both acBTX-treated and control rats, there was an inverse relationship between the quantal content of an endplate and its MEPP amplitude.5. The MEPP frequency of endplates from control muscles was positively correlated with the quantal content. However, this correlation was not found in aBTX-affected muscles.6. Three hours after a single injection of aBTX the amplitude of the MEPPs was reduced to about 60 % of control values but no increase of the quantal content was found. During the first few days of aBTX treatment the quantal content gradually increased; it reached a plateau between 20 and 30 days.7. The results suggest the existence of an adaptive mechanism, operating at individual endplates, in which retrograde signals at the motor nerve terminals modulate ACh release when neuromuscular transmission is endangered by block of acetylcholine receptors. MS 1036 J. J. PLOMP AND OTHERS

“…The amplitude of the m.e.p.c. in the presence of DEPP was not underestimated, since at a much higher concentration, 100 /LM, DEPP still has no curarizing action (Miledi, Molenaar & Polak, 1980), in contrast to neostigmine which at concentrations above 1 /LM depresses slightly the m.e.p.c. amplitude.…”

Section: Cholnnedtera8e Inhibitionmentioning

confidence: 99%

Acetylcholinesterase activity in intact and homogenized skeletal muscle of the frog.

Miledi¹,

Molenaar²,

Polak³

1984

Self Cite

SUMMARY1. Enzymatic hydrolysis of acetylcholine (ACh) was determined in intact frog sartorius muscles or their homogenates. The Vmax was 29 nmol min' in intact muscles and 46 nmol min' per muscle in homogenates, and the Km was 6 and 0-2 mM, respectively. The muscle was divided into small segments, which were homogenized; the junctional cholinesterase (ChE) accounted for 60 % of total enzyme activity.2. At low substrate concentrations the rate of hydrolysis was up to 30 times higher in homogenates than in intact muscles. This difference was greatly reduced at very high substrate concentrations. It appears that most of the ChE in intact muscle is 'occluded' to external ACh, mainly because the ChE at the edges of the synaptic cleft prevents the ACh from reaching the enzyme situated further inwards, which consequently does not contribute to its hydrolysis; homogenization makes all synaptic ChE accessible to added ACh.3. Incubation of sartorius muscles with collagenase caused an 80 % decrease in ChE activity (determined in homogenates) of end-plate-containing parts which became similar to that in end-plate-free parts on which collagenase had little effect.4. Histochemistry showed that the tendon-muscle junction contained folds which were stained intensively for ChE.5. Diethyldimethylpyrophosphonate, neostigmine, eserine, and di-isopropylfluorophosphonate inhibited ChE activity in this order of potency. The I50 values (i.e. the concentrations of the drugs which caused a 50 % inhibition) were about 5 times higher in intact than in homogenized tissue.6. Neostigmine, 0 15 and 0 4 /SM, increased the time constant of miniature end-plate currents 1-3-and 1-8-fold, and slowed down ChE activity of muscle homogenates by 1-4 and 2-1 times, respectively, without significantly affecting ACh hydrolysis by intact muscles. This indicates that synaptic ChE is not present in large excess. 7. It is concluded that ChE activity measured in homogenates presents a better t To whom reprint requests should be addressed.