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First published online 25 April 2006
doi: 10.1242/jcs.02931

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Endocytosis of GABA_B receptors modulates membrane excitability in the single-celled organism Paramecium

¹ Department for the Study of the Territory and its Resources (DIP.TE.RIS.), University of Genoa, Corso Europa 26, 16132 Genova, Italy
² Department of Communication, Computer and System Sciences (DIST), University of Genoa, Viale Causa 13, 16145 Genova, Italy
³ INFM and Department of Physics, University of Genoa, Via Dodecaneso 33, 16146 Genova, Italy
⁴ IFOM Center of Cell Oncology and Ultrastructure, Department of Experimental Medicine, University of Genoa, Medical School, Via de Toni 14, 16132 Genova, Italy
⁵ Department of Experimental Medicine, Section of Pharmacology and Toxicology, University of Genoa, Viale Cembriano 4, 16148 Genova, Italy
⁶ Institute of Biophysics, CNR Genoa, Via De Marini 6, 16149 Genova, Italy

^* Author for correspondence (e-mail: ramoino{at}dipteris.unige.it)

Accepted 10 February 2006

GABA_B receptors modulate swimming behavior in Paramecium by inhibiting dihydropyridine-sensitive Ca²⁺ channels via G-proteins. Prolonged occupancy of GABA_B receptors by baclofen results in a decrease in GABA_B receptor functions. Since changes in the number of cell-surface GABA_A 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 GABA_B receptors in P. primaurelia. In this study, we use immunostaining in electron and confocal microscopy to demonstrate that constitutive internalization of GABA_B receptors in P. primaurelia is mediated by clathrin-dependent and -independent endocytosis. Indeed, GABA_B 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 GABA_B receptor endocytosis into clathrin-coated and -uncoated vesicles represents an important mechanism in the modulation of swimming behavior in Paramecium.

Key words: GABA_B receptor, Endocytosis, Clathrin, Adaptin complex, Caveolin, Ciliated protozoa