Endocytosis of GABAB receptors modulates membrane excitability in the single-celled organism Paramecium.
GABAB receptors modulate swimming behavior in Paramecium by inhibiting dihydropyridine-sensitive Ca2+ channels via G-proteins. Prolonged occupancy of GABAB receptors by baclofen results in a decrease in GABAB receptor functions. Since changes in the number of cell-surface GABAA receptors have been postulated to be of importance in modulating inhibitory synaptic transmission in neurons, we have studied the cell-surface expression and maintenance of GABAB receptors in P. primaurelia. In this study, we use immunostaining in electron and confocal microscopy to demonstrate that constitutive internalization of GABAB receptors in P. primaurelia is mediated by clathrin-dependent and -independent endocytosis. Indeed, GABAB receptors colocalize with the adaptin complex AP2, which is implicated in the selective recruitment of integral membrane proteins to clathrin-coated vesicles, and with caveolin 1, which is associated with uncoated membrane invaginations. Furthermore, when endocytosis is blocked with hypertonic medium, cytosol acidification, filipin or with a peptide that disrupts the association between amphiphysin and dynamin, the effect of baclofen on swimming is increased. These results suggest that GABAB receptor endocytosis into clathrin-coated and -uncoated vesicles represents an important mechanism in the modulation of swimming behavior in Paramecium.
Ramoino
Paola
P
Department for the Study of the Territory and its Resources (DIP.TE.RIS.), University of Genoa, Corso Europa 26, 16132 Genova, Italy. ramoino@dipteris.unige.it
Gallus
Lorenzo
L
Beltrame
Francesco
F
Diaspro
Alberto
A
Fato
Marco
M
Rubini
Patrizia
P
Stigliani
Sara
S
Bonanno
Giambattista
G
Usai
Cesare
C
D000595:Amino Acid Sequence; D000818:Animals; D001418:Baclofen / Q000494:pharmacology; D022163:Clathrin-Coated Vesicles / Q000187:drug effects / Q000378:metabolism; D004705:Endocytosis / Q000187:drug effects / Q000502:physiology; D005455:Fluorescent Antibody Technique; D018755:GABA Agonists / Q000494:pharmacology; D058786:GABA-B Receptor Agonists; D018613:Microscopy, Confocal; D008969:Molecular Sequence Data; D010247:Paramecium / Q000187:drug effects / Q000378:metabolism / Q000502:physiology*; D018080:Receptors, GABA-B / Q000378:metabolism*
D018755:GABA Agonists; D058786:GABA-B Receptor Agonists; D018080:Receptors, GABA-B; D001418:Baclofen