ࡱ> 5745@ bjbj22 .&XXZ, !!!!!!$R EQE 0 p;4|.0   DEE$ Long-term exposure of recombinant GABAA receptors to neurosteroid dehydroepiandrosterone sulfate (DHEAS) Julija Erhardta, Maja Jazvinaak Jembrekb, Kety Mirkovi Kosb, Danka Peri ib and Dubravka `vob `tracb a Department of Animal Physiology, Faculty of Science, University of Zagreb, Zagreb, Croatia b Division of Molecular Medicine, Ruer Boakovi Institute, Zagreb, Croatia Long-term exposure to a variety of drugs including benzodiazepines, barbiturates and steroids induces different adaptive changes in GABAA receptors, the major fast inhibitory neurotransmitter receptors in the mammalian brain. In order to better understand the underlying mechanisms, human embryonic kidney (HEK) 293 cells stably expressing recombinant alb2g2S GABAA receptors (the most common type of GABAA receptors) were exposed for 24 and 48h to neurosteroid dehydroepiandrosterone sulfate (DHEAS). The aliquots of membrane preparations obtained from control and DHEAS-treated cells were used in the saturation binding studies with [3H]flunitrazepam and [3H]t-butylbicycloorthobenzoate ([3H]TBOB) to determine the possible changes in the binding parameters (Kd and Bmax), as well as in the functional interactions between GABAA receptor binding sites. Exposure of HEK 293 cells stably transfected with recombinant alb2g2S GABAA receptors to 100 mM DHEAS for 24 and 48h have not changed the maximum number nor the affinity of the binding sites for benzodiazepines and convulsants. Moreover, no significant differences between the groups were observed in LP$ & L N l n \ ^ ` h H   " wdwYwhoohoomH sH $hoohQw0JB*H*mH phsH !hoohQw0JB*mH phsH hoo0JB*phhoohoo0JB*H*phhoohoo0J>*B*H*phhoohoo0J>*B*phhoohoo0JB*ph$hoohoo0JB*H*mH phsH !hoohoo0JB*mH phsH h)0JB*mH phsH " ^ $dha$gd[- $dha$gdQw   /68rw/0]avw}ƵƙƵƙƆƆsƆƙ``Ƶ$hqNhQw0JB*H*mH phsH $hoohQw0J6B*mH phsH $hoohQw0JB*H*mH phsH hu0JB*mH phsH hhv0JB*mH phsH !hoohoo0JB*mH phsH !hoohQw0JB*mH phsH $hoohQw0JB*H*mH phsH )hoohQw0JB*OJQJmH phsH %"$&(*,0:<`bzn"͸͸{jW$hooh:0JB*H*mH phsH !hooh:0JB*mH phsH h:0JB*mH phsH Uhoo0JB*mH phsH !hoohoo0JB*mH phsH hhv0JB*mH phsH )hoohQw0JB*OJQJmH phsH $hoohQw0JB*H*mH phsH !hoohQw0JB*mH phsH hu0JB*mH phsH the potency (IC50) of DHEAS to modulate [3H]TBOB binding or in the efficacy (Emax) of GABA to potentiate [3H]flunitrazepam binding. These results have demonstrated that unlike benzodiazepines, chronic DHEAS treatment does not affect expression and functional coupling of GABAA receptor binding sites. If, as previously suggested, allosteric uncoupling is related to the development of functional and behavioral tolerance following prolonged treatment with GABAA receptor ligands (as in the case of benzodiazepines), then one might presume that chronic treatment with DHEAS will not produce tolerance. "$&RTv~$FHϼ૚tfUGU9GUhu0JB*mH phsH hoo0JB*mH phsH !hoohQw0JB*mH phsH hhv0JB*mH phsH $hhvh=Z0JB*H*mH phsH $hhvh=Z0JB*H*mH phsH !hhvhhv0JB*mH phsH !hhvh=Z0JB*mH phsH $hoohoo0JB*H*mH phsH !hoohoo0JB*mH phsH h:0JB*mH phsH !h:h:0JB*mH phsH &(:VZ^46\^   6 n z ϼϼ~!h[-hQw0JB*mH phsH h:0JB*mH phsH hu0JB*mH phsH !hoohQw0JB*mH phsH $hooh[-0JB*H*mH phsH !hooh[-0JB*mH phsH !h[-h[-0JB*mH phsH h[-0JB*mH phsH ,1h. A!"#$% @@@ NormalCJ_HaJmHsHtHDA@D Default Paragraph FontRiR  Table Normal4 l4a (k(No List8O8 Qwapple-style-spanBOB Qwapple-converted-spaceZ&ij/{|Y\000000000p/{|Y\O90X0 M90M90O90X "   8@0(  B S  ?\j./z\ |rw]a8;JUnr\\Dubravka `vob `trac  oo:qN?Ohv7uQw[-=Z)@LZPP P P@PPUnknownGz Times New Roman5Symbol3& z Arial" & &??!r4dVV3HX)?QwProlonged exposure to a variety of drugs, including benzodiazepines, barbiturates and steroids induces different adaptive changes in GABAA receptors often tolerance and dependenceLeoDubravka `vob `tracOh+'0 T`lx    Prolonged exposure to a variety of drugs, including benzodiazepines, barbiturates and steroids induces different adaptive changes in GABAA receptors often tolerance and dependencerolLeoeoeo Normal.dotxDubravka vob trac2brMicrosoft Word 10.0@@4|@4|?՜.+,0 hp|  |nVA Prolonged exposure to a variety of drugs, including benzodiazepines, barbiturates and steroids induces different adaptive changes in GABAA receptors often tolerance and dependence Title  !"#%&'()*+-./01236Root Entry FP1E4|8Data 1TableWordDocument.&SummaryInformation($DocumentSummaryInformation8,CompObjj  FMicrosoft Word Document MSWordDocWord.Document.89q