Inotropic effects of activation and pharmacological mechanisms in cardiac muscle

TIPS - November 1981 able. Compounds such as isebutylmethylxanthine, theophylline, and caffeine which distinguish between P site and R site adenosine receptors, are not .,,elective for R, and R, receptor subtypes, in analogy with other receptor systcms subtype selec-

tive antagonists would provide an important tool for the study of adcqosinc reccptots. The concept of subclasses of aaenosine receptors and thc development of subtype selective antagonists could have interesting implications for drug therapy. As an important example, theophylline can be mentioaed as one of the mainstays of therapy in acute bronchospasm and obstructive pulmonary diseases. Although inhibition of phosphodiesterase and alterations in intracellular calcium metabolism have long been considered as the basis of the therapeutic action of the methylxanthines, recent evidence demonstrates that an antagonistic interaction with adenosine receptors may contribute to the beneficial effects of theophylline s. At present, it is not known which subtype of adenosine receptot's may be involved in the broncholytic action of theophylline, but subtype selective antagonists could result in therapeutic advances of the type encountered in the pursuit of selective agonists and antagon-

3~~3 isis f o r other hormone and drug receptors. in conclusion, the first direct binding studies have provided a new approach to investigating adenosine receptors. It will Ix: interesting to see whether an extension of t h e ~ studies to other cellular systems and the development of further s u b t y w ~lec-

live radioligands will yield greater insights into the physiological and molecular prop-

I I Sth~.d~t lilt. ~

[

Kilt,

II

P,,h-t, m t

~u,l I ~..,,t I

4.7

I_" l)uIIa I~ told ~It-I ~I.~. • I ~ l'~"~I ! l'~m,-~,. ,4

I U' l i w , _'14. ~ . . .,~'_ I).d,, I '~ Nim.:ktll'~.t~,.,r~ 'l I'.,n. I l b , , . R l" "H,¢!11¢. J- anti %k, d n l c k P ~1'~'~,~) Plmrma, oh.lt~t . i 2 ~ 14 I~dll.tm,, %1 a . , I Rl..Ic,, ! '~ I I ' ~ I H t , . . / I't,,, t c , I t n l %q I tl' Bt,,I ~'~ I~.I'I I" ~Au P II I ' h d h , . I ~A, . ILdl,. K .~tl R m . d , t , I¢ I;

erties of adenosine receptors. Reading

list

1 % a t l i n . A. and R,dl. i. X~ ( IO7O1 t.l+,! l ' h , i , , l a , , d h. I :l- 23 2 I]urn,,tt~.-k. ( i 11'~;21 Pharnul,,,I R,'t 24 l l u r n , , h ~ k . ~ ~ I Iq,7*,t) Ifl Phl~t,,b,tl, al a n d R,'t,ulato O' Funct#om o f A d e m m n e a n d .4&'rune .%ucleotute~ IRacr. H. P. a n d D r u m m o n d . ( i I . cd~l.

pp. 3-32 4 ila,,huu. R..I and I ~nh.,n. I ~ I Itl72) I th' ~,t II. 1143-11 ¢,4 5 Frcdhohn.B. FI ( I t l ~ l l ) lrcml~ I'hurma,,,1% t 2.

I'r,,, Sail t , , , I ~,t I ~ I " ' . ~ ' ~ 1 " " ' ' l i " %tll~abt. ! mid It,,,! I I t'~,q ~,,,,~,; .~chmwd¢l~'rll~ ..Inh. P h a r m a k , I ", I ~. I "v 1~" I ~. ~llh.otw... M .lib| r i , , l t ~ I x i i,~,,~,l I',,,, x , t ,

, I."~ %i*,],',ChJ~ I~,,*,,Ir~ ~: n..',,;d. I' .m,! i,L,t,t.,,.

211 ~Rhq..lht. I . I I~tlt. R ,,,d I ~hI, l i i I Ill

tdliH' ~'~ HI ( t , / h

(I)rumm,,nd

(~

I. (,t.,

12t~- 132 h l.onth,,,. {'. a n d ~ ' o l i f . J. ( I tl'?7) Pr,~

%all .4< dd Sci. /.;..'~.,4. 74. "~4~2-54Sh 7 I.ondo,,,. ( ".. t,,o~-r. D M. F a n d ~,~,~dll. J. I I ,~.,,aq Proc..'GttlAcad..~ci. U.S..4. 7 7 . 2 5 f , I 25:.-,t V a n ( ' a l k c r . D.. Muller. M. a n d i l a m p r c ~ h t . B. ( I q 7 8 ) .%ature I L o n d , m l 27~. ,~ ~,~-,~4 I 9 Mall~m. ( . C . tlcrt. R. t ' . ,rod I=ain. J. \ . (I~Fr~) J. BioL ( h e m . 2 5 3 . 3 1 1 4 - 3 1 2 2 Ill Durra. P. a n d Mu',laia. %. J. ( I t~7t~) J. Ph, ar,m, ,,1 l-xp. lher. 211. 40e~-5Ol

t Irt~h _%hwahr t.~ P r o f i ' ~ , , r and ( ttmr-:an ,,t l'hur,r:.t.lo~lt a n d Ioin,,Io~:~ at tlw ( nt~crtttat B,,,rn tt:" re'( t'lgcd Ilt~ ,jt'~re(' It! ,v(dl, lilt" ~tr ] t#~t ~, ,t,t tilt [ ~1 t-t,r~ttar ( i o m n g e n and alter hahzhtatt,,n #,r t,h,ar,,~actdo~'t at lhg ianlt" tttll:'#rMfl lit" ~.tt'/|J" Ill lilt U,.dtytm~ hc it,,t h~t h M , tl, m , t , , , , r t,: l'~,~,~ [ , ,,,:

the ~att,,,hd Itt~ltt~,t~., ,,~ tic,/6~

~th.c,/~,,,;t/t. ,:,

ta, ~dtx at B~,nt~ :n / U - U

trophysiolog), biochcmistr~ and ion fluxcour kno~ ledge of individual pn~'cs.~ controlling calcium movement,,, and con,,:quentl) contractility,, has greati.~ increased 4ut'.nT.ts. None the It,,,,. the ~,a,*t amount of a~ailablc data h,t~c iloI led to a generall.~ accepted solution of the clcctromechanical coupling puzzle. Rather. the complcxit.~ of the problem, the quantit.~ ot available data. and the existence of ,,e,, oral conflicting lewd.,, toaard a solution ha~c spavined the development of a numlx'r ot different and to a con,,iderablc c,~tcnt irreconcilable modcN indeed, it ha.', become a full-time iob just to kccp up ~ ith and criticall.,, evaluate the a~ailablc data in P h a r m a k o l o g ~ c h e s b ~ r t u t der Universititt Tiibingen, Wilhelmstr. 5h. D- 7400 l'iibb~gen, b: R. (;. order to determine ~ hich model, a~,tx'ct,~of given models or modifications of given 'Staircase'phenomena in heart muscle sbnply reflect changt:~ o f the amount o f cah ium at its models arc mo,,t Iikch to Ix..correct. In thi,, release sites according to textbook explanatfims. A vailabh, evhtence, however, implicate.~ context the intcr~al-',trcnglh phenomena intracellular sodium and the sodium pump as well as ¢:vclic A M P and enzymes invoh'ed m of i~,olated heart muscle preparation,, regulatory protein phosphorvlation #~ potentiation phenomena. A better un&,rstanding o f remain perhap,, our mo~,t imi~rtant .,,ct ot these mechmtisms can be an important aid b~ the investigation o f cardioactive drags and guide tx~,,t,,. In tile almo,,t 20 .~car,, ,,mcc Koeh-\t,cm:r possibly will be relevant h! rehm'd areas o f n'search, such as elector-secretory coupling and and Blinks" revic~ ot the intcrval-,,trcngth neurophysiology. relationship T. the amount of literature For more than a century the complex con- decades major methodological advances v~hich has appeared on the subject is at tractile responses of isolated heart tissues have allowed multiple approaches to the least as great as that cited b~ those authors to changes of rhythm have been thought to problem of how membrane excitation is up to 1963. For obvkms rea.,ams of space. reflect important, basic processes underly- coupled to generation of contraction in this short article cannot be a comprehening cardiac contractility. In the last two heart. At the levels of uitrastructure, dec- sive review. In the fidlo~ing ~vc can onh

Inotropic effects of activation and pharmacological mechanisms in cardiac muscle

H. Joachim Mensing and Donald W. Hilgemann

y" [vl~,~-~u.-L:%orlh-tl,,llandBh,mcdv.'al Prv,,~ I'l?,l -le,'~ t, 14* ,~l !~lllttl o~,,1:$,t" ~,~

304

T I P S - N o v e m b e r l ~81

authors attempted to interpret all interval--slrength phenomena in several heart preparations in terms of NIEA and PlEA L'. Over the years, however, convincing evidence has accumulated for the exis. tence of more than two inotropic effects of activation, as well as contractile behaviors which do not fit the NIEAIPIEA schemes. Therefore, the terms NIEA and PlEA can be applied without reservation only in the context of the Blinks/Koch-Weser analysis method for mammalian atrium. Separate mechanisms can be differentiated in various ways, the simplest being the clear kinetic separation of two effects in the same muscte preparation. Also, certain effects accumulate or decay in strong dependence on the number of beats made (beat-dependent kinetics), whereas others display a definite time course, regardless of contractile activity. Of most interest to the readers of this journal is the possibility of Fig. I. Schematic diagram o f a patch o f mammalian heart sarcolemma; two junctional cisternae o f sarcoplasmk" pharmacologically 'dissecting' different reticulum (SR) with coupling 'feet'¶ and some other components thought to be involved in the processes discussed mechanisms, which implies the potential to in the text. Stud), left to right. Numbers i -4 at the arrows refer to Table I. Thick arrows repre~era calcium movements. differentiate and ultimately to identif~ celFcLsIs,.dium inw,,rd current Ls'a. causes action potential gAP) upstroke, which opens a hypothetical calcium lular points of drug action. Available evichannel in the "feet'; an out]low of SR-calcium via this rome during A P-overshoot" ,ocould corres~md to the "transient outward current'. 1~o,and perhaps resolve the enigma this current ha~ presented up to now=. Diffusion resis- dence strongly indicates the existence of at tance of the e.rterna~' lamina as (~;ellas laterally oscillating calciatn binding sites at the outer aspect of the plasma- least three different mechanisms of frelemma would favor an initial lateral spread o f the out/lowing calcium. In this model o f cation movemen~ (com. quency inotropy (increase of force with pare Re~ 5. 8-10), the calcium which activates contraction by binding to troponin enters the cell from the increase of frequency) with variable preglycoca#.x m an Cent/y/e~changeforNa, (representing an outward current, 'lc'ta), and also by slow inward current, dominance in different types of heart musIs~ via phospho~lawd 'slow channels'. S. C Phosphoo, lation by protein kinase, PK (namely, the cationic catalytic cle. Neither the variety of clearly separable sub.unit o f cAMP-dependent PKL is en/mnced during depolarization, while phosphoprotein phosphatase, PPPase. is active in;lependent o f the A P ~. Acti~ator calcium is nminly taken up by the SR (2Ca/A TP pump); cal. effects nor our increased knowledge of cell cium transport mec/mnisms in mitochondria and sarcolemma (.see right side o f the figure) may also contribute, and physiology is reflected in the terminology will take part in diastolic restoration o f the initial (or reswd) slaw o f the muscle, as does the 3Nal2KI I A TP pump. currently in use. For the sake o f clarin', other imoortant components and processes have not been included, such as Ca binding sites, We think the time has come to introduce potassium channels, adenylale cyclase, ~almodulin, and regulator), phosphor),lations other than that o f the slow a mechanistic terminology, which, along channel. with a neutral descriptive terminology, should promote a better conceptualization attempt to outline roughly the develop- Blinks and Koch-Weser, for example, and more adequate analysis of the procesments which have taken place, some prob- introduced the terms NIEA and PlEA ses which we want to understand. Ftmherlems which have arisen, and how we feel (negative and positive inotropic effects of more, if general terms are coined, they this work can be effectively advanced. As a activation) in connection with a careful, should be potentially useful in the descrippoint of reference for the reader, a mod- quantitative analysis of the inter- tion of electrophysiological findings= as ified versions.~°of Langer's 'one-way' con- val-strength relationship of m am m al i an well as activation-dependent effects in cept of calcium movements in mammalian atrium ~. According to this concept, each other excitable cells. For the time being we myocardiums (see also Ref. ~) is presented activation produces some PlEA and a suggest the following three main classificain Fig 1. This model integrates many find- greater amount of NIEA, which decay tions of interval-strength phenomena (cf. ings and could resolve some controversies. slowly between beats. At long intervals in Table !): effects of activation related to It has been shown to generate in detail the the range of minutes (rested state contrac- changes of ceflular calcium stores (Cainterval-strength phenomena of guinea pig tionsI, RSC), mammalian atrium develops related effects of activation: Ca-REA), atrium, as well as the effects and interac- strong contraction force; the NIEA repre- effects of activation related to cellular tions of several phar~:" ~ological agents on sents the process responsible for a fall of sodium (Na-REA), and effects of activathat relationships. steady-state force with increase of fie- tion related to regulatory protein phosqu,:mcy in a low frequency range, or within phorylation (PP-REA). just a few beats during continual stimulaA terminology for a multitude of effects tion after RSC. The PIEA represents the The development of an adequate ter- gradually cumulating process responsible Clmnges in cellular calcium stores minology for tli~: interval-strength for increase of force with increase of freNearly all inotropic effects reflect phenomena is still an unresolved and quency in a higher frequency range. Con- changes in calcium transients in the important problem. The terminology used traction strength equals RSC strength myofibrillar space. Determinants of these naturally reflects, and may also influence, minus cumulated NIEA plus cumulated transients include fill levels of cellular calour understanding of the phenomena. PIEA. Apart from their analysis, these cium stores, membrane permeabilities to

305

T I P S - N o v e m b e r ! g81

calcium, and kinetics of calcium binding and pumping. Under the term "calciumrelated effects of activation" (('a-REA) we would include only those after-effects of activation which result from deviations of the fill levels of calcium stores from their equilibrium at the rested state. The group of Ca-REA to be de~ribed here are prominent in mammalian myocardium, but are minor or absent in frog heart t-tT.u, which lacks a welldeveloped sarcoplasmic reticulum (SR)"u. Present concepts u . " about SR-Ca-REA can be summarized as follows. At excitation, calcium is released from the junctional cistemae of the SR and in part is responsible for contractile activation. For relaxation most calcium is pumped into longitudinal SR, and SR release sites are replenished with a time constant of ~ m e w h a t less than one second (delayed restitution"). Individual contraction cycles may result in either a net decrease or increase of SR-calcium. Thus, force transients for resumption of muscle stimulation after various rest periods in atrium very typically begin with a beat-dependent negative staircase (NIEA cumulation); in ventricular preparations the corresponding beat-dependent staircases are often positive in direction (an SR-Ca-related mechanism of frequency inotropy). When SR-calcium is reduced below its rested state level, calcium is resupplied quite slowly (NIEA decay; time constant of around 0.5 rain). If SR-calcium is increased above its rested state level, it is lost similarly slowly in the absence of contractions. Most surplus SR-calcium can however be lost in a single beat (e.g. decay of postextrasystolic potentiation) s'~s. This is a good argument in favor of a depolarization-dependent outflow of calcium TM from the SR to the cell surface (glycocalyx), from where only part reenters the cell and contributes to activations,~e. In such a one-way concept of calcium movements a large part of activator calcium leaves the cell at the very next beat, which explains strongly beatdependent kinetics of SR-Ca-REA. It should be mentioned at least that the function of other cellular calcium stores may show up in certain kinetic analyses of activation-dependent effects. Mitochondriai buffering of cytosolic calcium and rapid transients of cell surface calcium may be the domina,:i Ca-REA mechanisms in frog heart. Changes in cellular sodium Cytosolic sodium (Na,) strongly influences cardiac contractility4`*''. Recent find-

ings from numerous laboratories indicate an elcctrogenic, potential-dependent sarcolemmal 3-Na-for-I-Ca exchange, which during positive displacement of the membrane potential will promote influx of extracellular calcium in exchange for Na,. correlated to the third power of Na,. The resulting outward current opix~c5 ~imultancously flowing inward currents and may coniplicatc voltage-clamp analysis m.n. During diastole this exchange carrier coordinates active outward transport of Ca and Na by the respective sareolcmmal ion pumps. Especially with hmg action potentials. Na-influx via 'fast' and "slow"channels may produce a slow frequen~-related increase of Na,. and consequently enhance Ca-

influx w r activation in exchange for Na,U . ' . At least in guinea pig ventricular myocardium this slow "Na,-REA" tyl~." o| frequency" inotropy plays a pronounced role j'. The effect can accumulate in Ca-free medium and depends on extraccllular .~xlium. it is diminished or enhanced by druKs which reduce or increa~ Na influx (e.g. tetrodotoxin, veratridinep ". The time constant of approach to a new stead~ state (one to ~vcral minutes) is slowed by inhibition of outward sodium transporl with heart glycosides". This suggests the possibility to quantitatively cstip lie inhibition or stimulation of the sou, .. pump in simple functional experiments on the appropriate music preparation.

Developed Force 30

(mN)

f n

20

tl

\, /

,== ==

h

_

~.,, - -..-..~ -~6

10

.o..G." .. /

0

II

0

l

I

I

t

l 0.5

'

'

i

i

I

,

1.0

' (Hz)

'

'

• 1.5

Stimulation Frequency Fig. 2. Automated .frequency-fim'e curves jh, m the le~ atrzun, o f a 22¢)g gmnea pig" 1.2 m *t t'u ! Krehs-tlenseleu solution I 15 m,tt dextrose I low temperature ¢280(9 to reduce rested state t ontracuon.~ for t~olanon of tht" PLEA. Peak developed force wider isometric conditions ~s recorded directly aga6gt .~ti,mlatitm frequent'). ~h~ch i.~ increased and decreased exponentiall) in each loop ¢]'=foe ± t~" k =0.003 sec ~L)s.6.Arrows indicate the direction o f automatic frequency change. Carres 1.2, -] and 4 show the O'pical effect o f [3.adrenergic stimulation by on'i prenaline (control. 10 -T, 4 x 10 -T and 1.6 × t O-e ;t, respectively): accumulation o f PIEA is shifted to a lower frequency range, and at the higlwst concemruzion there £~a subslanaal increase o f the ,ested stale contracuon. ('urr~ 4 to 5 demonstrate the rerersal ofthe effect o]o~iprenaline by carbachol fcurre 5:1.0 x ! 0 ~' .toorcqTrenaline + 6 x !0-" M carbachol). The effect ofnifedi~ine, demonstrated in curves 4 to ~. is simply to scale down du relalions h i p with no effect on the frequency range ,;f P I E A accumulatkm ¢curve ~, 1.0 x 10-" .u orciprenalme + 5 x i 0 -" ,~!nifedipine ).

306 Regulatory protein phosphorylation O n the basis of the following findings Robison et al.~S suggested in 1965, that an enzyme or e n z y m e system regulated by c A M P might be involved in the production of PLEA. ( ! ) The positive inotropic effect of moderate adrenergic stimulation in mammalian atrium reflects mainly an increased production of P l E A per beat'. Since the atrial P l E A decays with a time constant in the o r d e r of a minute, it cumulates roug0-,iy in pro F ,'rtion to heart rate ~, and adrenergic inotropy shows a corresponding frequency dependence'. (2) Biochemical work implicated c A M P to be the second messenger of most or all effects of B-adrenergic stimulation in hearP s. (3) The production of cAMP, itself, does not depend on ccmtractile activity. B-adrenergic stimulation most probably exerts its positive inotropic effect by enhancing calcium inward current via socalled slow channels. Presumably, the phosphorylation of slow channels or a related sarcolemmal protein by c A M P dependent protein kinase results in the availability of a greater n u m b e r of channels at activation '7. The hypothesis o f Robison et ai. might therefore mean that 'slow channel" phosphorylation is (partially) activation-dependent s, due either an effect of depolarization on protein kinase or on the substrate itself*. Changes of the limiting slow channel conductance would be expected to parallel accumulation and decay of the artrial PLEA, the rate of which would be determined by phosphoprotein phosphatase activity. Although a direct testing of this hypothesis seems nearly impossib~,, at present, a thorough pharmacological analysis of the interval-strength relationship o f guinea pig atrium strongly supports this hypothesis over available alternatives s. When slow inward current is restricted by one o f several means (blockade by nifedipine, action potential shortening by adenosine, or reduction of the extracellular calcium), the positive inotropic effects o f

77PS - N o v e m b e r 1981 TABLE I. Sumn)aryof main inotropic effectsof activation: mechanisticterminology in relation to ,~sme descriptive terms: g)me characteristicsof individualeffects and remarks on possible interpretation and occurrence in different types of heart muscle, Numbers I--4 refer to corresponding processesin Fig. I. Mechanistic clarification

Descriptiveterms

Characteristics (v = time constant)

Commentary

SR-Ca-RFA

(Delayed) Restitution~t

vin theorderof several hundred milliseconds. Beat-dependentcumulation/ decay; slow decay in the absenceof contractions, Tin the order of 0.5 miu, quite variablen. depends on preparation, conditions

Ca translocationwithin the SR to release sites (junctional cistcrnae) ( I ). Net change of SR-calcium;fast and large efflux/influxduring activation; in the absence of contractions slow loss (2a) of surplus Ca, or gain (2b) in the case of SR-Ca deficit (e.g. NIEA of an atrial rested state contraction).

¢ in the order of one t' to ~veraP minutes, reflects Na-transportactivity(3). is slowed by heart glyc~)sidesTM

Prominent in guinea pig ventricleTM. In principle, Nai-REA could be positive. negativeor absent, depending whether an activation results in a net Na+gain. loss or no change.

(Short) 'Staircase'/ last pha~ m~ Post-extrasystolic potentiation NIEA~ Nat-REA

'Frequency inotropy/ potentiation'; 'staircase'/slow phase (guinea pig ventricle)'+

SIowchanncl 'Frequencyinotropyl v in the order of a minute PP-REA potentiation'; in mammali;matriums (Qt0 'staircase': 2.7)s, much faster in frog (atrial) PLEA" atrium even at lower temperature~a.t+:in mammalianventricle, ~"might well be of the order of seconds at physiological temperatures*.

Predominant in mammalianatrium: effect produced per beat is small (short action potential) but cumulates strongb because of slow decay. In preparations with long action potentials decay might be faster (phosphoprotein phosphatase activity(4) higher)to counteract a greater production of effect per beat.

*A paraile' ",mdationof SR-Ca-REA and slowchannet PP-REA with fast time constant in mammalian vem~triclec, ~ excluded.

/3-adrenergic stimulation and increased frequency approach assymptotically the same submaximal force level, which theoretically would correspond to maximal slow channel phosphorylation. /3-adrenergic stimulation, as well as phosphodiesterase inhibition or cAMP analogues, shift the accumulation of frequency inotropy to a lower frequency range. This effect can be antagonized for the most part by carbachol, which is known to reduce e n h a n c e d c A M P levels. Decreasing slog' inward current by the cAMPindependent means m e n t i o n e d above, however, reduces the maximum of accumulation with little or no effect on the frequency range o f accumulation; see Fig. 2. Thus, with strong/3-adrenergic stimulation antagonized by a reduction of slow "The mole,cular mechanism of such an effect is a inward current, the normal mechanism of matter of speculation; the following could happen. frequency inotropy in guinea pig atrium With positive displacement . f the membrane can be nearly abolishedU; presumably potential a negatively charge0 part of the slow slow channel phosphorylation approaches channel (a gate?) moves out somewhat from the its m a x i m u m at a very low frequency, inward-facing protein surface and in con.~quence can be phosphoq,'lated mort readily by protein and no other mechanism of frequency kinase. Vice versa, a positivdy charged cytosolic inotropy is operative. molecule, like the catalytic subunit of cAMPU p to now a positive correlation bedependent protein kinase, will be attracted to the tween frequency inotropy and slow inward inner aspect of the sarcolemma when its outside current has been r e p o r t e d only for frog becomes negatively charged at excitation. This attraction is Ifighly dependexa on the charge the atrium '+. Earlier evidence from another molecule bea,s, and will be strongest at thin group implicated a phosphorylation penetrating sarcolemmal prol,dns with high dielec- mechanism in this effect'*. In mammalian tric constant, which the slow ,:~aanael may well be. atrium the crucial electrophysiological

experiments are still lacking. In ve~tricular m y o c a r d i u m o f mammals, where ionic currents have been intensively investigated, the slow inward current transient regularly decreases with increase of frequency. Either the slow channel P P - R E A is insignificant in mammalian ventricle, or it is masked behind another more pronounced effect. If P P - R E A kinetics in ventricle were m o r e rapid than in atrium, due to a higher phosphoprotein phosphatase activity, it would be difficult to separate P P - R E A and SR-Ca-REA. It should be kept in mind that calcium/calmodulin-dependent protein kinase is likely to be involved in generation of certain after-effects of activation, be it at the plasma m e m b r a n e or elsewhere in the cell. In mammalian myocardium stimulation o f the SR-calcium p u m p is a prime candidate 's.

Outlook The analysis of interval-strength phenomena deserves careful attention in the context of rapid progress being made currently in the study of excitationcontraction coupling and mechanisms of drug action in the heart. It may be assumed that every or almost every important process controlling cardiac contractility will be represented in some interval-strength p h e n o m e n o n or in the contraction curve

TIPS

-

N o v e m b e r i 981

itself, Drug effects on the.~ p r o c e s ~ s can be studied in experiments with quite simple, inexpensive equipment on relatively intact and undisturbed myocardium preparations. Thus, m a n y aspects of cellular and subcellular handling of calcium and ~ d i u m may be investigated without the use of radioactive tracers, cell fractionation, biochemistry, electrophysiology or other d e m a n d i n g methods. Functiomd experiments on muscle strips and data analysis can be economically performed by application of a u t o m a t e d e q u i p m c n P and microca~mputer technology. A 'fingerprint" of a drug's action could be obtained in parallel experiments on several preparations, e.g. at1 ial strips and papillary muscles from the same heart. With the advent of a reliable and adequate mathematical model of excitation-contraction coupling it will become possible to perform a semiquantitative analysis o f drug effects on distinct cellular processes in 'simple' functional experiments. Reading list ! Antoni, H., Jacob. R. and Kaufmann. R. (I g~9) Pfldgers Arch. 3lib, 33-57 2 Blinks, J. R. and Koch-Weser. J. ( 1t~61) J. Pharmacol. Exp. Ther. 134, 373-389 3 Boyen, M. R. and Jewell. B. R. (19811)Prog. Biophys. Mol. Biol. 36, !-52 4 Fozzard, H. A. (19771 Ann. Rev. Physiol. 39. 201-220 5 Hilgemann,D. W. ( 198111New Perspective~on the lmervaI.Strength Relatimt, hip of Guinea Pig Atrimn. Ph.D Thesis. Faculty of Biology.University of Tiibingen, F.R.G. 6 Hilgemann. D.. Englert, R. and Mensing. H. J. (I g77) Experientia 33, 1629-163 I 7 Koch-Weser, J. and Blinks, J. R. (Itlh3) Pharmacol. Rev. ! 5, 61)I---652 8 Langer, G. A. (19741 in The Mammalian Myocardium CLanger,G. A. and Brady. A. J. eds) pp. 193-217, John Wiley and Sons, New York, London, Sidney and Torcmto 9 Langer, G. A., Frank, J. S. and Brady, A.J. ( 1976) b~t. Rev. PhysioL 9, 191-237 Ill Mensing, H. J. (1979) Naunyn-St'hmicdeberg.~ Arch. Pharmakol. 308, R 35 I I Morad, M. and Goldman, Y. ( It~731 Prog. Biophys. Mol. Biol. 27, 257-313 12 Mullins, L. J. (1979) Am. J. PhysioL 23h. CI()3--Ci I(I 13 Niedergerke, R. and Page, S. (1977) Prec. R. Soc. Lond. Ser. B It~7,333-362 14 Noble, S. and ~rhimoni,Y. ( 1981)J. Physiol. 310, 57-~)5 15 Robison, G. A., Butcher, R. W., O~c, I.. Morgan. H. E. and Sutherland, E. W. (19¢~5)Mol. PharmacoL i, I hS-177 16 Seibel, K. (1979) Naanyn-Schmiedebergx Arch. Pharmakoi. 308, R 6 17 Tsien, R. W. ( 1977lAdy (:relic NucleotMeRes. 8. 363-4211 18 Walsh, M. P., LePeuch, C. J.. Vallet, B., Cavadote, J.-C. and Demaille.J. G. ( i 980) J. Mol. ('ell. CardioL !2, 11191-1Hit 19 Wohlfart, B. (1979) Acta PhysioL Stand. II~. 395-4119

3117

Heinz J t m c h t m Mett;ing graduated m m e d u u z e at d . " University o f Marburg in 1970. A t the d¢l~trtments o f

pharmacology m Marburgan,} lt~bmgen he engaged m the ,levelopnwm and impm:',,ment o] meth,,d~ ~,r nu,a~urement of re~pirator~ membolt~m and }iuu uon of Lsolated heart intrude, brelmg enth~ia.stt, about basic m~t~('leresearch, he has been contrihtal~g m u ~ h of his spare nine to a criticaletaltuuum of aata f r o m the hterature with a foctt~ on the ~nergctics and pharmacology of e~cimtion-comracnm~ coupling.//e obtained ht~ M.D. at the Universtty of fubingen m I U77.

I b m I h l g c m a m , t~ a n u n v e o f b , s a ti,. { , , m p h . t , d m, ,~t , [ hti undergradtatn" ;tudte~ m httd, ~ and th ; ,r°'n~ ,d medu uu'. a~ ~ ¢// as a Pit D m pharu . . . . . . / , ~ : t t , I o , ' O t at the Unlter~tt~ ,,f [ubmk.~'n !1¢ has hc~.n trcat ng t~,,lawd guinea p,,, atria re, drue~, p,~L~¢m~ a n d .trantlc stlmldus palterrt~ fl~r aheml s i t ~,ars ~l~t~'r ~J po~t~b~ctond fl'll,,~ ihtp at the ( entre ,h" R,,clu.tchc lf.'er,,ll International..~itrashourg. h," L~ pursuing que~rt.,n~ ,t electro-me~ lhlnl(al ColtphsZ~, at thc. ( ardlottz~(td, zr Re~earch Laboratoru's ,,] the [ n ~ c r tt~ ~,f ( .~ht,,rm, at l.os .4ngeh'~

Discovery of a n t i t u m o r agents f r o m natural sources Matthew Suffness and John D. Douros Natural Products Branch l h v l x m n o f ( ~lnt-~.r I tea,men,. R o o m ~ 14. Bhur Bad, fine..~.h ~ ~( , ,1~.~~dlr R, ,a,t. ~d~ cr

Spring. MD 20010. U.S.A.

Whih, various f o l k h w e remedies fi~r d w trcauncnt o]'canccr /tare a 1,mg iU.~torv, the m ~th'rn d e v e l o p m e n t o f anticancer drugs f r o m natural sourte.~ ¢phmts, microbes, marine ani,:al~ ) i.~ a relatively n e w area o f research. The complexity and difficulty o f drug deteh~pmet:t as it relates to cancer is probabh'. ,lOt generalh', lolder.~lood and we wi.~h first to (ii.~tus.~;.some ~~./" the p r o b l e m s and partial solutions bt'fi~re mentioning briefly xome o f t ! I f ('t,mp(~totd.~ di.xcovered a n d the outlook fi)r the fitmn'. In vivo s c r e e n i n g - t h e m o d e l s a n d their sensitivity

There are well over I tl(I types of human cancer and it is quite u n l i k e h that a m one screening model ~,ould be predictive fi~r clinical activity in yen' m a r e tumol types. it is clearly recognized in clinical thcrap.,, that most if not ncarl~ all clinicalh uscft,! agents have a narrow spectrum of use and that this depends not onh, on the organ site of the tumor but on the specific t~pe of tissue from which it is derived. Since human tumors are so diverse and so specifically sensitive it is quite understandable that tumors of o t h e r mammalian species (mice, rats) may not adequately reflect what substam,-es would be effective in man. In order to have reproducible test systems, rodent tumors were initially t h o r n fi)r screening.

Th¢,,c ~ere ,,table cell line,, capable of being readil.~ transplanted and ha~ing rapid gro~ th eharacteri.,,tic.,, so that ,,cree,ing experiments could bc oomph:ted in 20-3(I da~ s. in recent .~cars ,do~ er gro~ ing rodent tumor~ ha~e I~:en tl~ed as modcl~ fl~r the sloxs gr~,,,,r~ solid tumors m m a n

s~hich are most rc.,i, ;.atlt 1o chemotherapy. hut these model,, • uffer from being nonhuman in origin and trom being ~,table line., instead of spomaneow, [onlols. |'h¢ room recent development has Ix'on the u~" of the athvmic mouse ~hich is immunodcficient and ~ill accept transplants of human tumors. Con.~quently, much of the current screening can be done u.qng human t u m o r models. A review by Goldin et al. * on m vivo screening for anticancer agents coscrs this topic well. l; ~4.,%1¢r Nt*llh Ho[|,lt)d Br,,r~|cd)~.*' I~r,*-', tU~,l