Patch-clamp-induced perturbations of [Ca(2+)](i) activity in somatotropes.
Somatotropes and GC cells, a GH-producing cell line, exhibit [Ca(2+)](i) oscillations that result from rhythmic Ca(2+) action potentials. Determination of this operating mode required simultaneous recording of both parameters by fura-2 imaging and patch-clamp techniques. In order to test whether patch recording induces artificial alteration of the [Ca(2+)](i) oscillatory pattern, we recorded separately or simultaneously [Ca(2+)](i) and membrane potential. In the absence of any other stimulation, seal formation in patch-clamp recording evoked by itself a 2.5- to 4-fold persistent increase in basal [Ca(2+)](i), speeded up their frequency (from 0.03-0.17 to 0.4 Hz) and changed their pattern to a tonic mode. Patch-induced [Ca(2+)](i) increase was reproduced by mechanical contact between the pipette and the membrane. It was reduced by nifedipine, a blocker of L-type Ca(2+) channels, as well as by removal of external Na(+). It was fully blocked by external Ca(2+) removal or gadolinium. All patch-clamp-induced perturbations were reversed by membrane hyperpolarization. We propose that patch-clamp recording evokes Ca(2+) entry through L-type Ca(2+) channels either directly, or indirectly via membrane depolarization. This shows that patch recordings in endocrine cells showing mechanosensitivity have to be interpreted with caution, and explains why long-lasting patch recordings are so difficult to obtain.
Robert
C
C
Dynamique des Systèmes Neuroendocriniens, INSERM U159, Paris, France.
Tseeb
V
V
Kordon
C
C
Hammond
C
C
D000818:Animals; D001683:Biological Clocks; D002118:Calcium / Q000378:metabolism*; D002121:Calcium Channel Blockers / Q000494:pharmacology; D020746:Calcium Channels, L-Type / Q000502:physiology*; D002478:Cells, Cultured; D005456:Fluorescent Dyes; D016257:Fura-2 / Q000031:analogs & derivatives; D005682:Gadolinium / Q000494:pharmacology; D007987:Gonadotropin-Releasing Hormone / Q000494:pharmacology; D008297:Male; D009543:Nifedipine / Q000494:pharmacology; D018408:Patch-Clamp Techniques; D010902:Pituitary Gland / Q000166:cytology* / Q000187:drug effects / Q000502:physiology*; D051381:Rats; D017208:Rats, Wistar; D013314:Stress, Mechanical
D002121:Calcium Channel Blockers; D020746:Calcium Channels, L-Type; D005456:Fluorescent Dyes; C049925:fura-2-am; D007987:Gonadotropin-Releasing Hormone; D005682:Gadolinium; D009543:Nifedipine; D002118:Calcium; D016257:Fura-2