Gene/Protein Disease Symptom Drug Enzyme Compound
Pivot Concepts:   Target Concepts:
Query: EC:3.4.24.3 (collagenase)
 18,340 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

We used the loose patch voltage clamp technique and rhodamine-conjugated alpha-bungarotoxin to study the regulation of Na channel (NaCh) and acetylcholine receptor (AChR) distribution on dissociated adult skeletal muscle fibers in culture. The aggregate of AChRs and NaChs normally found in the postsynaptic membrane of these cells gradually fragmented and dispersed from the synaptic region after several days in culture. This dispersal was the result of the collagenase treatment used to dissociate the cells, suggesting that a factor associated with the extracellular matrix was responsible for maintaining the high concentration of AchRs and NaChs at the neuromuscular junction. We tested whether the basal lamina protein agrin, which has been shown to induce the aggregation of AChRs on embryonic myotubes, could similarly influence the distribution of NaChs. By following identified fibers, we found that agrin accelerated both the fragmentation of the endplate AChR cluster into smaller patches as well as the appearance of new AChR clusters away from the endplate. AChR patches which were fragments of the original endplate retained a high density of NaChs, but no new NaCh hotspots were found elsewhere on the fiber, including sites of newly formed AChR clusters. The results are consistent with the hypothesis that extracellular signals regulate the distribution of AChRs and NaChs on skeletal muscle fibers. While agrin probably serves this function for the AChR, it does not appear to play a role in the regulation of the NaCh distribution.
 ...
PMID:Effect of agrin on the distribution of acetylcholine receptors and sodium channels on adult skeletal muscle fibers in culture. 165 12

The present study evaluates suitable in vitro methods for the assessment of the inhibiting properties of four principally different antidepressant drugs. This was done by comparing the acute effects of antidepressants on autonomic receptor binding (homogenates) together with parallel tests evaluating the biological activities of the receptor systems in collagenase-isolated rat parotid acini. The responses were measured as receptor-activated changes in cyclic nucleotide formation and acinar oxygen consumption. Muscarinic acetylcholine receptor binding, carbachol-induced cGMP formation, and oxygen consumption all reflected the various inhibiting effects of the antidepressants tested. Measurements of the carbachol-induced O2 consumption was however, the most sensitive method and may be considered a well-suited and reliable parameter concerning the expected severity of anticholinergic side-effects caused by medication. The disturbing 'dry mouth' symptoms following treatment with amitriptyline or mianserin are however, also attributed to their substantial adrenoceptor-blocking effects, which are best demonstrated by alpha 1-adrenoceptor binding studies in combination with measurements of the adrenaline-induced O2 consumption in the rat parotid gland.
 ...
PMID:In vitro methods for the assessment of the inhibitory effects of antidepressants in rat parotid glands. 166 35

Ascorbic acid is the active component of fetal brain extract that induces increased acetylcholine receptor (AChR) expression in L5 rat clonal muscle cell cultures. The induction of AChR expression, as determined by 125I-alpha-bungarotoxin binding, occurs with a delay of 20-25 h. We report that the delayed increase in AChR can be triggered by a 5-h exposure to ascorbic acid. These studies suggest that intermediary processes may be involved. Ascorbic acid treatment also causes a threefold increase in collagen secretion in L5 cultures by 3 h. The rapid increase in collagen secretion and the delayed induction of surface AChR suggested that there may be a link between these two responses. However, although bacterial collagenase eliminates secreted collagen, it had no effect on the increase in surface AChR. Thus, the ascorbic acid effect on elevating AChR expression is independent of its effect on collagen secretion.
 ...
PMID:Acetylcholine receptor regulation in L5 muscle cells is independent of increases in collagen secretion induced by ascorbic acid. 196 67

Cultures of rat myotubes from 18-day-old embryos produce both globular (G) and asymmetric (A) forms of acetylcholinesterase (AChE; EC 3.1.1.7), mostly G1, G4, and A12 and a small proportion of A8. We show that all forms are partly intracellular and partly exposed to the extracellular medium; the A forms and their intra- and extracellular distribution are not modified when myotubes are grown in the presence of spinal cord neurons. In these cocultures, however, AChE patches may be detected immunohistochemically at sites of neuromuscular contacts. These patches represent a very minor proportion of AChE activity. We found that collagenase removes AChE patches but not the acetylcholine receptor clusters with which they coincide. This digestion specifically decreases the level of the A12 form. cis-Hydroxyproline, an inhibitor of collagen synthesis, reduces the level of G1 and blocks the synthesis of A forms.
 ...
PMID:Acetylcholinesterase in cocultures of rat myotubes and spinal cord neurons: effects of collagenase and cis-hydroxyproline on molecular forms, intra- and extracellular distribution, and formation of patches at neuromuscular contacts. 215 53

The present study demonstrates the effects of the antidepressant, amitriptyline, and the acetylcholine antagonist, atropine, on the stimulation-induced rise in cytosolic, free Ca2+ (Cai2+). The changes in Cai2+ of collagenase-isolated rat parotid acini were measured by means of the Ca2(+)-sensitive dye, fura-2. It was found that stimulation by carbachol resulted in a maximal increase of 582 +/- 34 nM (mean +/- S.E.) in Cai2+ with a ks of 5.8 +/- 1.3 microM. Adrenaline caused a rise of 380 +/- 22 nM in Cai2+ with a ks of 0.5 +/- 0.2 microM. Amitriptyline and atropine were found to inhibit the carbachol-induced rise in Cai2+ with dissociation constants (kI) of 105 and 1.25 nM, respectively, in the absence of agonist. The adrenergic-induced rise in Cai2+ was inhibited by amitriptyline with a kI of 45 nM. Amitriptyline was found to inhibit both receptor classes by a competitive or mixed type of inhibition. Similarly, atropine exerted the same type of inhibition on the acetylcholine receptor. Amitriptyline and atropine were found to be mutually exclusive for competing for substrate binding on the receptor. These findings are consistent with a common binding site for amitriptyline and atropine on the acetylcholine receptor, possibly in close proximity with, but different from the substrate binding site. The stimulation-induced cell shrinkage evoked by the loss of electrolytes and water from the acini was measured by a 90 degree light scattering signal. It was found that this method makes possible the detection of autonomic side-effects of antidepressants on acini suspended in protein-containing media.
 ...
PMID:Inhibitory effects of amitriptyline on the stimulation-induced Ca2+ increase in parotid acini. 234 Aug 55

Functional acetylcholine receptor (AChR) and sodium channels were expressed in the membrane of Xenopus laevis oocytes following injection with poly(A)+-mRNA extracted from denervated rat leg muscle. Whole-cell currents, activated by acetylcholine or by depolarizing voltage steps had properties comparable to those observed in rat muscle. Oocytes injected with specific mRNA, transcribed from cDNA templates and coding for the AChR of Torpedo electric organ, expressed functional AChR channels at a much higher density. Single-channel currents were recorded from the oocyte plasma membrane following removal of the follicle cell layer and the vitelline membrane from the oocyte. The follicle cell layer was removed enzymatically with collagenase. The vitelline membrane was removed either mechanically after briefly exposing the oocyte to a hypertonic solution, or by enzyme treatment with pronase. Stretch activated (s.a.) currents were observed in most recordings from cell-attached patches obtained with standard patch pipettes. S.a.-currents were evoked by negative or positive pressure (greater than or equal to 5 mbar) applied to the inside of the pipette, and were observed in both normal and mRNA injected oocytes indicating that they are endogenous to the oocyte membrane. The s.a.-channels are cation selective and their conductance is 28 pS in normal frog Ringer's solution (20 +/- 1 degree C). Their gating is voltage dependent, and their open probability increases toward more positive membrane potentials. The density of s.a.-channels is estimated to be 0.5-2 channels per micron 2 of oocyte plasma membrane. In cell-attached patches s.a.-currents are observed much less frequently when current measurement is restricted to smaller patches of 3-5 micron 2 area using thick-walled pipettes with narrow tips. In outside-out patches s.a.-currents occur much less frequently than in cell-attached or inside-out patches. AChR-channel and sodium channel currents were observed only in a minority of patches from oocytes injected with poly(A)+-mRNA from rat muscle. AChR-channel currents were seen in all patches of oocytes injected with specific mRNA coding for Torpedo AChR. In normal frog Ringer's solution (20 +/- 2 degrees C) the conductance of implanted rat muscle AChR-channels was 38 pS and that of sodium channels 20 pS. The conductance of implanted Torpedo AChR channels was 40 pS. The conductance of implanted channels was similar in cell-attached and in cell-free patches.(ABSTRACT TRUNCATED AT 400 WORDS)
 ...
PMID:Patch clamp measurements on Xenopus laevis oocytes: currents through endogenous channels and implanted acetylcholine receptor and sodium channels. 243 68

The concentration-dependent actions of neostigmine, a carbamate anticholinesterase agent, were studied on the acetylcholine receptor channel complex in voltage-clamped twitch fibers of costocutaneous muscles of garter snakes. Low concentrations of neostigmine (10(-6) or 10(-5) M) increased miniature endplate current (MEPC) amplitude and the time constant of MEPC decay without changing the relationship between the MEPC decay time constant and membrane potential. Acetylcholine- or carbachol-induced endplate current fluctuation spectra were well fitted by a single Lorentzian curve with a characteristic frequency and single-channel conductance unaltered by low concentrations of neostigmine. Concentrations of neostigmine greater than 5 X 10(-5) M decreased MEPC amplitude and split the decay of MEPCs into two components, one faster and one slower than the control rate. These effects were both voltage and concentration dependent. Spectra of current fluctuations recorded in concentrations greater than or equal to 5 X 10(-5) M neostigmine required two time constants, one faster and one slower than the control. Two component spectra were also obtained with carbachol-induced current fluctuation spectra, indicating that these effects of neostigmine were direct and not a consequence of acetylcholinesterase inhibition. Similar results were also obtained in muscles pretreated with collagenase to remove junctional acetylcholinesterase. The fast and slow time constants obtained from current fluctuation spectra decreased and increased, respectively, with either increases in the concentration of neostigmine or membrane hyperpolarization when analyzed in the same fiber. The effects of neostigmine on channel lifetime were reversible with washing. These results indicate that the effects of neostigmine are concentration dependent. Concentrations greater than 2.5 X 10(-5) M exhibit direct effects on the endplate receptor channel complex which are unrelated to acetylcholinesterase inhibition. These actions include: a prolongation of the gating kinetics of the endplate receptor channel complex, the production of an altered state of the receptor channel complex evidenced by a high frequency component to current fluctuation spectra, and a direct action to block the acetylcholine receptor.
 ...
PMID:Concentration-dependent effects of neostigmine on the endplate acetylcholine receptor channel complex. 257 18

Coculturing of rat embryonic muscle cells with spinal cord explants resulted in the formation of large numbers of acetylcholine receptor aggregates on the myotube surface, compared to those found on muscle cells grown in the absence of nervous tissue. Remarkably fewer receptor aggregates were formed when, upon addition of nerve explants, these cocultures were treated with either cis-hydroxyproline (a specific inhibitor of collagen production) or collagenase. The possibility is raised that collagen participates in the aggregation of acetylcholine receptors and in synapse formation.
 ...
PMID:Aggregation of acetylcholine receptors in nerve-muscle cocultures is decreased by inhibitors of collagen production. 629 Sep 48

The existence of nicotinic acetylcholine receptors (AChRs) on the motor nerve terminals of vertebrates has long been controversial. We have re-examined this issue by electron microscope autoradiography with [125I] alpha-bungarotoxin, following separation of nerve terminals from muscle fibers by collagenase and protease treatment. We found no label over nerve terminal membranes other than that due to background, and we calculate upper limits of less than 0.1% of the postsynaptic AChR density for nerve terminals in frogs, lizards, and mice. We conclude that there are essentially no presynaptic acetylcholine receptors that bind alpha-bungarotoxin at vertebrate neuromuscular junctions.
 ...
PMID:Absence of [125I] alpha-bungarotoxin binding to motor nerve terminals of frog, lizard and mouse muscle. 682 66

The role of collagen macromolecules in the aggregation of acetylcholine receptors on the surface of cultured rat muscle cells was investigated. Both the synthesis and secretion of collagen, as well as acetylcholine receptor aggregation were stimulated by treatment of muscle cultures with an embryonic brain extract. Treatments with cis-hydroxyproline or monensin, which interfere with collagen synthesis and secretion, significantly reduced the number of acetylcholine receptor aggregates detected. Incubation of the cultures with embryonic brain extract induced a decrease in extractability of acetylcholine receptors. This effect could be reversed by treating the cells with bacterial collagenase. In addition, affinity purified antibodies against collagen of types I and IV inhibited the aggregation of acetylcholine receptors by stimulated brain extract. These data suggest that neurotrophic factor(s) present in embryonic nerve tissue have the ability to induce collagen production; the collagen formed plays a role in the formation and/or stabilization of acetylcholine receptor aggregates and possibly also of synaptic connections.
 ...
PMID:Involvement of collagen in the aggregation of acetylcholine receptors on cultured muscle cells. 713 Jan 74


1 2 Next >>