Cycl i c 3' , 5' -AMP Rel ay i n Di ct yost el i um di scoi deum V. Adapt at i on of t he CAMP Si gnal i ng Response dur i ng c AMP St i mul at i on ABSTRACT In Di ct yost el i um di scoi deum, ext r acel l ul ar c AMP act i vat es adenyl at e cycl ase, whi ch l eads to an i ncr ease in i nt r acel l ul ar CAMP and t he r at e of c AMP secr et i on . The si gnal i ng r esponse to a const ant CAMP st i mul us is t er m i nat ed af t er sever al m i nut es by an adapt at i on mechani sm . The t i me- cour se of adapt at i on to st i mul i of 10-6 or 10 -7 MCAMP was assessed . We used a per f usi on t echni que to del i ver def i ned c AMP st i mul i to [ 3 H] adenosi ne- l abel ed amoebae and moni t ored t hei r secr et i on of [ 3 H] c AMP. Amoebae wer e pr et r eat ed with 10 -6 or 10 -7 M c AMP f or per i ods of 0 . 33- 12 m i nut es, and t hen i mmedi at el y gi ven t est st i mul i of 10 -8 Mto 2 .5 X 10 -7 Mc AMP. The r esponse to a gi ven t est st i mul us was pr ogr essi vel y at t enuat ed and f i nal l y ext i ngui shed as t he dur at i on of t he pr et r eat ment st i mul us i ncr eased . Dur i ng pr et r eat ment with 10 - 6 MCAMP, t he r at es of at t enuat i on coul d be r anked accor di ng to t he concent r at i on of t he t est st i mul us . The r esponses to t est st i mul i of 10 - 8 , 5 X 10 - 8 , 10 -7 , or 2 .5 X 10 -7 Mc AMP wer e ext i ngui shed af t er - 1, 2 . 25, 2 . 5, and 10 m i n, r espect i vel y . 1 .5 m in of st i mul at i on with 10 -7 MCAMP was necessar y to ext i ngui sh t he r esponse of a t est st i mul us of 10 - 8 M c AMP. Our dat a suggest t hat adapt at i on begi ns wi t hi n 20 s of st i mul at i on, r i ses r api dl y f or -2 .5 m i n, and r eaches a pl at eau af t er - 10 m in . The absol ut e r at e of rise was f ast er dur i ng pr et r eat ment with 10 - e t han wi t h 10 -7 M CAMP. These r esul t s suppor t a wor ki ng hypot hesi s in whi ch t he occupancy of sur f ace c AMP r ecept or s l eads to changes in t wo opposi ng cel l ul ar pr ocesses, exci t at i on and adapt at i on, t hat cont r ol t he act i vi t y of D . di scoi deum adenyl at e cycl ase . The aggr egat i on of Di ct yost el i um di scoi deum is medi at ed by pr opagat ed si gnal s of cycl i c adenosi ne 3' , 5' -monophosphat e ( cAMP) . The si gnal i ng r esponse has been st udi ed bi ochem i- cal l y ( 5, 13, 14) . The bi ndi ng of c AMP to cel l sur f ace r ecept or s ( 7, 8) l eads to t he act i vat i on of adenyl at e cycl ase ( 5, 12) , an i ncr ease in t he l evel of i nt r acel l ul ar CAMP, and secr et i on of t he newl y- synt hesi zed c AMP ( 5) . If t he st i mul us is wi t hdrawn dur i ng t he r esponse, si gnal i ng prompt l y subsi des . Si gnal i ng r esponses al so t er m i nat e spont aneousl y af t er a f ew m i nut es of sust ai ned st i mul at i on as a r esul t of an adapt at i on pr ocess ( 3) . The mechani sm of adapt at i on appear s to i nvol ve a r ever si bl e, t i me-dependent decr ease in sensi t i vi t y to c AMP st i mul i . Ser i al i ncrement s in t he l evel of ext r acel l ul ar c AMP evoke successi ve si gnal i ng r esponses f ol l owed by adapt at i on to each new stim- ul us concent r at i on . Duri ng aggr egat i on, adapt at i on pr esum- abl y r egul at es t he magni t ude and dur at i on of c AMP si gnal i ng 554 MARY C . DI NAUER, THEODORE L . STECK, and PETER N. DEVREOTES Depart ment s of Bi ochem i st r y and Medi ci ne, The Uni ver si t y of Chi cago, Chi cago, I l l i noi s 60637 r esponses and pr event s amoebae f r om r espondi ng per pet ual l y to t hei r own secr et i ons . We have pr evi ousl y shown t hat adapt at i on begi ns to i ncr ease bef or e t he si gnal i ng r esponse t er m i nat es, per si st s af t er t he decl i ne of t he r esponse, and decays with f i r s t - or der ki net i cs as soon as t he CAMP st i mul us is r emoved ( 3, 6) . In t hi s i nvest i - gat i on, we sought to def i ne t he t i me- cour se f or t he rise in adapt at i on upon t he i nt r oduct i on of a c AMP st i mul us . As a wor ki ng hypot hesi s, we assumed t hat a st i mul us, per cei ved as an i ncr ease in t he f r act i onal occupancy of cel l sur f ace c AMP r ecept or s, pr oduces changes in bot h an exci t at i on pr ocess t hat l eads to act i vat i on of adenyl at e cycl ase, and an opposi ng pr ocess, adapt at i on, t hat bl ocks such act i vat i on ( 6) . If t hi s hypot hesi s is val i d, t he l evel of adapt at i on m i ght be det ect ed at any time dur i ng t he si gnal i ng r esponse by i nt er - r upt i ng t he st i mul us and subst i t ut i ng a t est st i mul us of l ower Ti t s J OURNAL OF CELL BI OLOGYzyxwvutsrqponmlkjihgfedcbaZYXWV " Vot ut i +E 86 AUGUST 1980 554- 561 ©The Rockef el l er Uni ver si t y Pr ess - 0021- 9525/ 80/ 08/ 0554/ 08 $1 . 00 on May 6, 2016 jcb.rupress.org Downloaded from Published August 1, 1980