Release of acetylcholine from embryonic myocytes in Xenopus cell cultures

Abstract

Acetylcholine (ACh) is important as the transmitter responsible for neuromuscular transmission. Here we report the non‐quantal release of ACh from embryonic myocytes. Co‐cultures of spinal neurons and myotomal muscle cells were prepared from 1‐day‐old Xenopus embryos. Single channel currents were recorded in the non‐innervated myocytes. When the patch pipette was filled with Ringer solution alone, spontaneous single channel currents occurred, which were inhibited by d‐tubocurarine (d‐Tc). The channel conductance appearing in Ringer solution (37.3 pS) was similar to that of an embryonic‐type ACh channel (36.9 pS), indicating that ACh is probably released from myocytes in normal Ringer solution. When the patch pipette was filled with anticholinesterase alone to prevent hydrolysis of ACh released from myocytes, both physostigmine and neostigmine in a concentration‐dependent manner increased channel open probability; it was reduced by d‐Tc or α‐bungarotoxin. Vesamicol and quinacrine, vesicular transporter inhibitors, reduced the channel open probability caused by ACh released from myocytes in the presence of neostigmine or physostigmine. Intracellular alkalinization with NH4Cl inhibited the ACh release from myocytes, whereas, extracellular alkalinization, brought about by replacing normal Ringer solution, with pH 8.6 Ringer solution enhanced ACh release. The immunocytochemistry of choline acetyltransferase (ChAT) showed that ChAT exists in both myocytes and neuronal cells but not in fibroblasts. These results suggest that embryonic myocytes are capable of synthesizing and releasing ACh in a non‐quantal manner. Extracellular alkalinization enhanced and intracellular alkalinization inhibited ACh release from myocytes.

Presence of a non-neuronal cholinergic system and occurrence of up- and down-regulation in expression of M2 muscarinic acetylcholine receptors: new aspects of importance regarding Achilles tendon tendinosis (tendinopathy)

“…The lack of further information is a drawback as (1) fibroblasts and myofibroblasts, i.e., counterparts of tendon tenocytes but occurring in other parts of the body, have been considered to be targets for acetylcholine (ACh; e.g., Jacobi et al 2002;Sekhon et al 2002) and (2) activation of ACh can be related to nociception and anti-nociception (see Dussor et al 2004). Furthermore, non-neuronal cells may produce ACh (Fu et al 1998;Fritz et al 2001), and a non-neuronal intrinsic cholinergic system has been described for certain vascular beds (e.g., Ikeda et al 1994;Haberberger et al , 2002Kirkpatrick et al 2003). Of interest in this respect are our recent findings that a non-neuronal cholinergic system is present in human patellar tendons, particularly in those exhibiting tendinosis (Danielson et al 2006).…”

Expression of the High-affinity Choline Transporter CHT1 in Rat and Human Arteries

“…However, there are several reports documenting the ability of muscle cells to synthesize ACh, either via ChAT or via the enzyme carnitine acetyltransferase. ACh-synthesizing activity and/or ACh release have been demonstrated in myoblasts (Krause et al 1995;Fu et al 1998), skeletal muscle fiber (Tucek 1982;Miledi et al 1982), and smooth muscle fibers of the human skin . Moreover, airway smooth muscle cells have been shown to display ChAT immunoreactivity .…”

Motoneurons of the axial swimming muscles in hatchlingXenopus tadpoles: Features, distribution, and central synapses

“…An attempt was made to mark the whole population of motoneurons by using an antibody to choline acetyl transferase. Because Fu et al (1998) have recently shown that such antibodies will mark isolated Xenopus spinal neurons in culture, it is probable that there is an antibody access problem in more intact tissue. Despite these qualifications, the sample is large enough to reveal overall trends and give a broad picture of the features and distribution of motoneurons, their output synapses and input connections.…”