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N-type, omega-conotoxin-sensitive Ca2+ channels mediate electrically evoked release of ACh in guinea pig trachea
D G Baker1, H F Don, J K Brown
1Pulmonary and Critical Care Medicine Section, Veterans Affairs Medical Center, San Francisco, California 94121.
Abstract:
To determine whether N- or L-type Ca2+ channels mediate acetylcholine (ACh) release from airway parasympathetic nerve endings, we compared the effects of omega-conotoxin (N-type inhibitor) and nifedipine (L-type inhibitor) on electrically evoked release of ACh in guinea pig trachea. Reconnected segments of guinea pig trachea were mounted in organ baths containing Krebs-Henseleit buffer, indomethacin (10 microM) to inhibit cyclooxygenase, neostigmine (1 microM) to inhibit acetylcholinesterase, and atropine (0.3 microM) to inhibit muscarinic autoreceptors, as well as phentolamine and propranolol to inhibit adrenergic receptors. After electrical field stimulation (EFS), aliquots of buffer were removed, and ACh was measured directly by high-performance liquid chromatography with electrochemical detection. Tracheas were stimulated for 10-min periods at a frequency of 5 Hz, and ACh release was measured for five separate periods (S1-S5) after treatment with increasing concentrations of omega-conotoxin or vehicle alone (acetic acid). Thirty minutes was allowed between stimulation periods. We found that EFS-evoked release of ACh was inhibited by omega-conotoxin in a concentration-dependent manner [mean effective concentration (EC50) approximately 8 nM] but was unaffected by vehicle treatment. In other experiments, ACh release was measured for two separate periods (S1 and S2), and between periods tracheas were treated with omega-conotoxin (1 microM), nifedipine (10-100 microM), tetrodotoxin (TTX), or buffer containing low (0.8 mM) Ca2+. ACh release was 12 +/- 2 (mean +/- SE) and 3 +/- 0.3 pmol.mg protein-1.min-1 before and during omega-conotoxin (n = 5, P < 0.05).(ABSTRACT TRUNCATED AT 250 WORDS)
Insights
N-type calcium channels, not L-type, mediate acetylcholine release from airway parasympathetic nerves. Omega-conotoxin significantly inhibited release, while nifedipine had no effect, confirming N-type channel involvement in neurotransmission.
Area of Science:
- Neuroscience
- Pharmacology
- Physiology
Background:
- Airway parasympathetic nerves regulate bronchomotor tone.
- Acetylcholine (ACh) is a key neurotransmitter released by these nerves.
- The specific calcium (Ca2+) channels involved in ACh release are not fully understood.
Purpose of the Study:
- To investigate whether N- or L-type Ca2+ channels mediate ACh release from guinea pig airway parasympathetic nerve endings.
- To determine the role of specific Ca2+ channel subtypes in cholinergic neurotransmission in the airways.
Main Methods:
- Experiments utilized isolated guinea pig trachea segments.
- Electrically evoked ACh release was measured using high-performance liquid chromatography.
- The effects of N-type (omega-conotoxin) and L-type (nifedipine) Ca2+ channel inhibitors were assessed.
Main Results:
- Omega-conotoxin inhibited electrically evoked ACh release in a concentration-dependent manner (EC50 ≈ 8 nM).
- Nifedipine did not affect ACh release.
- Tetrodotoxin and low Ca2+ also reduced ACh release, consistent with nerve-mediated release.
Conclusions:
- N-type Ca2+ channels are primarily responsible for mediating ACh release from airway parasympathetic nerve endings.
- L-type Ca2+ channels do not play a significant role in this process.
- Targeting N-type Ca2+ channels could be a therapeutic strategy for airway-related disorders.