A characterization of muscarinic receptor-mediated intracellular Ca2+ mobilization in cultured rat hippocampal neurones.
1. The properties of muscarinic receptor-mediated Ca2+ mobilization were investigated in hippocampal cultures using fluorescent imaging techniques. 2. Somatic responses to carbachol (1-10 microM) were observed in 21 % of neurones under control conditions (5.4 mM K+, 1. 8 mM Ca2+, 0.5-1 microM tetrodotoxin). Smaller responses were observed in Ca2+-free medium. 3. In cells where responses to carbachol were absent under control conditions, responses were often observed following depolarization with high extracellular K+ (16. 2-25 mM). These responses decreased in magnitude with time after the depolarizing episode. Mobilization of Ca2+ from stores using caffeine (50 mM) exhibited similar properties. 4. Carbachol responses were greatly facilitated in the presence of moderate elevations in extracellular K+ or Ca2+ levels (2- or 3-fold, respectively). These conditions were usually, but not always, associated with a small increase in cytosolic Ca2+ levels (< 50 nM). 5. Muscarinic responses in 10.8 mM K+ were inhibited by 80-95 % in the presence of the L-type voltage-gated Ca2+ channel antagonists nitrendipine (2-5 microM) or nifedipine (10 microM). Depletion of intracellular Ca2+ stores with thapsigargin (2-10 microM) blocked responses. 6. Oscillatory Ca2+ mobilizing responses were observed in some cells. Their expression was facilitated by moderate cytosolic Ca2+ elevations and by increasing the duration of carbachol exposure. 7. Ca2+ mobilizing responses were also observed in dendritic regions. These were smaller than somatic responses, but had faster decay kinetics. 8. In conclusion, muscarinic receptor-mediated Ca2+ mobilization in cultured hippocampal neurones shows a strong Ca2+ dependence. Moderate intracellular Ca2+ rises greatly facilitate muscarinic responses and uncover, in some cells, oscillatory Ca2+ mobilization. These effects appear to reflect the loading state of intracellular Ca2+ stores.
Department of Biomedical Sciences, Institute of Medical Sciences, University of Aberdeen, Foresterhill, Aberdeen AB25 2ZD, UK. a.j.irving@abdn.ac.uk
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AJ
Irving
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Collingridge
D000818:Animals; D002118:Calcium / Q000378:metabolism*; D015220:Calcium Channels / Q000502:physiology; D020746:Calcium Channels, L-Type; D002217:Carbachol / Q000494:pharmacology; D002412:Cations / Q000378:metabolism; D002478:Cells, Cultured; D018679:Cholinergic Agonists / Q000494:pharmacology; D004791:Enzyme Inhibitors / Q000494:pharmacology; D006624:Hippocampus / Q000166:cytology; D008564:Membrane Potentials / Q000187:drug effects / Q000502:physiology; D008856:Microscopy, Fluorescence; D009419:Nerve Tissue Proteins / Q000502:physiology; D009474:Neurons / Q000737:chemistry* / Q000187:drug effects / Q000378:metabolism; D010507:Periodicity; D011188:Potassium / Q000494:pharmacology; D051381:Rats; D011976:Receptors, Muscarinic / Q000502:physiology*; D019284:Thapsigargin / Q000494:pharmacology
D015220:Calcium Channels; D020746:Calcium Channels, L-Type; D002412:Cations; D018679:Cholinergic Agonists; D004791:Enzyme Inhibitors; D009419:Nerve Tissue Proteins; D011976:Receptors, Muscarinic; D019284:Thapsigargin; D002217:Carbachol; D011188:Potassium; D002118:Calcium