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ment of cocaine addiction. Because partialagonists or antagonists have different ef-fects, depending on the conditions, theymay be used as substitutes for abuseddrugs to facilitate abstinence, while alsoserving a prophylactic role by antagoniz-ing the effects of the abused drug in pa-tients that relapse. The partial opioidreceptor agonist buprenorphine is ex-pected to gain approval for use in US clin-ics as an alternative to methadone forheroin addiction. Ultimately, partial ago-nists also may be available for the treat-ment of cocaine addiction, and it seemsthat the D-3 receptor may be a viable tar-get. In the meantime, basic scientists arechallenged with the task of identifyingand validating the effects of partialdopamine agonists in animal models ofcocaine addiction, and determiningwhether such effects may be attributed toactions at the D-3 receptor or to otherdopamine receptor subtypes.

1. Pilla, M. et al. Selective inhibition of cocaine-seek-ing behaviour by a partial dopamine D-3 receptoragonist. Nature 400, 371–375 (1999).

2. Koob, G. F. & Le Moal, M. Drug abuse: hedonichomeostatic dysregulation. Science 278, 52–58(1997).

3. Roberts, D. C., Corcoran, M. E. & Fibiger, H. C.On the role of ascending catecholaminergic sys-tems in intravenous self-administration of co-caine. Pharmacol. Biochem. Behav. 6, 615–620(1977).

4. Caine, S. B. & Koob, G. F. Effects of mesolimbicdopamine depletion on responding maintainedby cocaine and food. J. Exp. Anal. Behav. 61,213–221 (1994).

5. Koob, G. F. Hedonic valence, dopamine, and mo-tivation. Mol. Psychiatry 1, 186–189 (1996).

6. Ahmed, S. H. & Koob, G. F. Transition from mod-erate to excessive drug intake: a change in hedo-nic set point. Science 282, 298–300 (1998).

7. Markou, A. & Koob, G. F. Postcocaine anhedonia:an animal model of cocaine withdrawal.Neuropsychopharmacology 4, 17–26 (1991).

8. Caine, .B. & Koob G.F. Modulation of cocaine self-administration in the rat through D-3 dopaminereceptors. Science 260, 1814–1816 (1993)

9. Caine, S. B. & Koob, G. F. Pretreatment with thedopamine agonist 7-OH-DPAT shifts the cocaineself-administration dose-effect function to the leftunder different schedules in the rat. Behav.Pharmacol. 6, 333–347 (1995)

10. Caine, S.B. et al. D3 receptor test in vitro predictsdecreased cocaine self-administration in rats.Neuroreport 8, 2373–2377 (1997)

11. Lamas, X., Negus, S. S., Nader, M. A. & Mello, N.K. Effects of the putative dopamine D-3 receptoragonist 7-OH-DPAT in rhesus monkeys trained todiscriminate cocaine from saline.Psychopharmacology 124, 306–314 (1996)

12. Nader, M. A. & Mach, R. H. Self-administration ofthe dopamine D-3 agonist 7-OH-DPAT in rhesusmonkeys is modified by prior cocaine exposure.Psychopharmacology 125, 13–22 (1996)

13. Spealman, R.D. Dopamine D3 receptor agonistspartially reproduce the discriminative stimulus ef-fects of cocaine in squirrel monkeys. J. Pharmacol.Exp. Ther. 278, 1128–1137 (1996).

Dept of Neuropharmacology, CVN-71The Scripps Research Institute

10550 N. Torrey Pines Rd

La Jolla, California 92037, USA

Email: gkoob@scripps.edu

Department of Psychiatry2Harvard Medical School

Alcohol and Drug Abuse Research Center

McLean Hospital, 115 Mill Street

Belmont, Massachusetts 02478, USA

NEWS & VIEWS

In stroke, complement will get you nowhereThe complement cascade has been proposed as another participant in the inflammatory events leading to neuraltissue damage after stroke. But only time, and many more experiments, will tell whether molecules designed to

inhibit specific pathways of the inflammatory response will pay off.

GREGORY J. DEL ZOPPOMUCH OF THE damage done to neuraltissue after stroke, or interruption

of blood flow to the brain, is the result ofinflammatory events. Very little isknown about the inflammatory mecha-nisms that lead to the destruction of neu-rons after cerebral ischemia. Onepathway that may be involved is thecomplement system. Complement is ahost defense system, made up of plasmaproteins which identify pathogens andinjured cells, recruit inflammatory cellsand induce cell lysis (Fig. 1).Complement components and specificcomplement receptors have been associ-ated with central nervous system inflam-matory disorders including Alzheimerdisease and multiple sclerosis1–3, yet thereis little information on a role for comple-ment in acute brain ischemia.Complement could also cause the in-creased mortality seen in recent inter-ventional stroke studies testing theplasminogen activator streptokinase—streptokinase may combine with circu-lating antibodies to activate thecomplement cascade. A paper by Huanget al. published in the 23 July issue ofScience4 suggests a role for complement inneuronal damage after experimental

focal brain ischemia.In the normal immune response, C1q,

the initiating factor in the classical com-plement cascade, binds to the surface offoreign cells to target them for destruc-tion (Fig. 1). Huang et al. examinedbrain tissue sections taken from a mousemodel of middle cerebral artery (MCA)occlusion, and observed that neurons,but not microvessels, in ischemic re-gions stained positively for C1q. The au-thors concluded that complementactivation was involved in neuronal in-jury after ischemia. Experiments in non-human primates have also shown thatthe endothelial cell–leukocyte adhesionreceptors P-selectin, ICAM-1, and E-se-lectin appear on microvascular endothe-lial cells following ischemic injury5,6.These molecules induce the accumula-tion of platelets, polymorphonuclear(PMN) leukocytes, and fibrin in the mi-crovasculature very early after ischemiaonset7,8 (Fig. 1). Murine adhesion recep-tor knockout experiments, and studiesusing rodent models in which these re-ceptors have been blocked pharmaco-

logically, provide further evidence forinflammatory cell involvement in ex-perimental stroke models.

To reduce inflammatory damage tocerebral tissue after brain ischemia,Huang et al. used a bifunctional mole-cule designed to inhibit both comple-ment activation and selectin-mediatedadhesive events. Previous studies havesuggested that sCR1, an inhibitor ofcomplement activation, can decreaseleukocyte accumulation in experimen-tal brain trauma9. Huang et al. used aform of sCR1 that was modified by sialylLewis x (sLex) glycosylation. The sLexmoiety binds cell surface selectins andblocks selectin - mediated cellular adhe-sion10. Administration of this hybridmolecule, sCR1sLex, to mice after is-chemic onset decreased the volume ofinjured cerebral tissue and reportedly re-duced neurological deficit for up to 23hours after MCA occlusion4. The au-thors claimed that the sCR1sLex mole-cule was localized to both C1q -expressing neurons and to microvesselsin the ischemic territory.

The exact mechanism by whichsCR1sLex reduces cerebral infarct size isnot clear from these experiments. The

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NEWS NEWS & VIEWS

authors suggest that sCR1sLex inhibitsactivation of the complement cascade, aswell as leukocyte and platelet recruit-ment to protect against neuronal injury.Presumably, the sCR1 portion ofsCR1sLex binds to the components ofthe multi-subunit C3 of C5 convertase inthe complement cascade (Fig. 1),whereas the sLex portion inhibits inflam-matory cell recruitment10. The extent towhich neurons express selectins or com-plement components after ischemic in-jury, however, remains to be determined.

This work raises several issues regard-ing the role of the complement system incerebral ischemic injury. Complementactivation occurs by three pathways, theclassical, lectin and alternative pathways(Fig. 1). Inflammatory processes involv-ing leukocyte adhesion receptor expres-sion on endothelial cells may upregulatecomponents of the complement sys-tem11. It is not known, however, whetheror not cells within the brain produce allthe components of the classical path-way12, or whether the lectin or alterna-tive pathways are also involved. Certaincomplement components and/or theirreceptors have been found on astrocytes,microglia, and neurons in both chronicand acute inflammatory conditions13. It

has not yet been de-termined whetherC1q is synthesized bythe neurons them-selves, or is producedby other cells andspecifically targets is-chemic neurons. Itwill also be importantto learn more aboutthe time frame atwhich C1q binds theneurons: is the com-plement pathway anearly response, or is itparticipating in thefinal stages of an al-ready dying cell?

Several technicalpoints about themeans of focal is-chemia inductionused in these experi-ments should be con-sidered. The period ofMCA occlusion is rela-tively short in contrastto other stroke modelsystems. Studies haveshown that after short(30 minutes) periods

of MCA occlusion, the development ofthe ischemic injury and the infarctionzone is not complete by 24 hours ofreperfusion, the time point at whichHuang et al. evaluated injury14,15.Therefore, it is likely that differences be-tween the treatment groups may havenarrowed or even disappeared by a latertime point. Additional controls shouldalso be done before we can consider thetherapeutic value of this approach.Mutant forms of the bifunctional agentshould be tested, such as molecules withaltered sLex (to prevent selectin bind-ing), a nonfunctional sCR1 portion (toaddress complement participation),along with a molecule containing bothinactive sCR1 and sLex. Other inhibitorsof selectin binding or complement acti-vation should also be tested. Answeringthese questions would provide more de-tailed information about the mecha-nisms by which sCR1sLex reducescerebral injury.

The exact events that occur during theearly phases of acute stroke are not wellunderstood, and a careful investigationof the role(s) of complement in brain in-jury following acute ischemia is a timelyventure. However, given the large num-ber of positive reports from other inter-

ventional approaches in rodent modelsof cerebral ischemia, it will be necessaryto gain better insight into the mecha-nisms of sCR1sLex protection, and to testthis approach in other models before thisapproach should be considered as a ther-apy for human stroke.

1. Reid, D.M., Perry, V.H., Andersson, P.B. & Gordon,S. Mitosis and apoptosis of microglia in vivo in-duced by an anti-CR3 antibody which crosses theblood-brain barrier. Neuroscience 56, 529–533(1993).

2. Yasojima, K., Schwab, C., McGeer, E.G. & McGeer,P.L. Up-regulated production and activation of thecomplement system in Alzheimer’s disease brain.Am. J. Pathol. 154, 927–936 (1999).

3. Compston, D.A. et al. Immunocytochemical local-ization of the terminal complement complex inmultiple sclerosis. Neuropathol. Appl. Neurobiol. 15,307–316 (1989).

4. Huang, J. et al. Neuronal protection in stroke by ansLex-glycosylated complement inhibitory protein.Science 285, 595–599 (1999).

5. Okada Y. et al. P-selectin and intercellular adhesionmolecule-1 expression after focal brain ischemiaand reperfusion. Stroke 25, 202–211 (1994).

6. Haring, H-P., Berg, E.L., Tsurushita, N., Tagaya, M.& del Zoppo, G.J. E-selectin appears in non-is-chemic tissue during experimental focal cerebral is-chemia. Stroke 27, 1386–1391 (1996).

7. Del Zoppo, G.J., Schmid-Schinbein, G.W., Mori, E.,Copeland, B.R. & Chang, C.-M.Polymorphonuclear leukocytes occlude capillariesfollowing middle cerebral artery occlusion andreperfusion in baboons. Stroke 22, 1276–1283(1991).

8. Okada, Y., Copeland, B.K., Tung, M-M. & delZoppo, G.J. Fibrin contributes to microvascular ob-structions and parenchymal changes during earlyfocal cerebral ischemia and reperfusion. Stroke 25,1847–1853 (1994).

9. Kaczorowski, S.L., Schiding, J.K., Toth, C.A. &Kochanek, P. M. Effect of soluble complement re-ceptor-1 on neutrophil accumulation after trau-matic brain injury in rats. J. Cereb. Blood Flow Metab.15, 860–864 (1995).

10. Rittershaus, C.W. et al. Recombinant glycoproteinsthat inhibit complement activation and also bindthe selectin adhesion molecules. J. Biol. Chem. 274,11237–11244 (1999).

11. Kato H., Kogure K., Liu X.H., Araki T. & Itoyama Y.Progressive expression of immunomolecules on ac-tivated microglia and invading leukocytes followingfocal cerebral ischemia in the rat. Brain Res. 734,203–212 (1999).

12. Afagh, A., Cummings, B.J., Cribbs, D.H., Cotman,C.W. & Tenner, A.J. Localization and cell associa-tion of C1q in Alzheimer’s disease brain. Exp.Neurol. 138, 22–32 (1996).

13. Gasque, P., Singhrao, S.K., Neal, J.W., Gotze, O. &Morgan, B.P. Expression of the receptor for com-plement C5a (CD88) is up-regulated on reactive as-trocytes, microglia, and endothelial cells in theinflamed human central nervous system. Am. J.Pathol. 150, 31–41 (1997).

14. Du, C., Hu R., Csernansky, C.A., Hsu, C.Y. & Choi,D.W. Very delayed infarction after mild focal cere-bral ischemia: A role for apoptosis? J. Cereb. BloodFlow Metab. 16, 195–201 (1996).

15. Endres M. et al. Attenuation of delayed neuronaldeath after mild focal ischemia in mice by inhibitionof the Caspase family. J. Cereb. Blood Flow Metab.18, 238–247 (1998).

Dept. of Molecular and Experimental Medicine

The Scripps Research Institute

10550 North Torrey Pines Road, MEM 132

La Jolla, California 92037, USA

Email: grgdlzop@hermes.scripps.edu

Fig. 1 Model of the complement cascade. Early components in theclassical, lectin or alternative pathways recognize injured cells and ini-tiate a cascade leading to inflammatory cell infiltration and activation,and destruction of the damaged cells. After C1q, mannose bindingligand (MBL) or C3b recognize and bind an injured cell, a proteolyticsignaling cascade is initiated. This cascade leads to generation of C3convertases that activate cleavage of C3 into C3a and C3b, generat-ing C5 convertase. C5 convertase activates cleavage of C5 into C5aand C5b. C3a and C5a attract basophils and eosinophils to damagedtissue. C5a also induces infiltration of PMN leukocytes, monocytesand macrophages to sites of injury. In the central nervous system, mi-croglia may be similarly activated by complement. Generation of theattack complex C5b-9 is also responsible for lytis of selected cell pop-ulations, such as bacteria and other cells.

Alternativepathway

(C3b)n

sCR1Basophils,

Eosinophils

*Phagocytosis

Cell lysis

Inflammatorycell infiltrationand activation

C5b-9

MBLMASP 1,2C4C2

C1q

C3

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C3b*

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C5-convertase

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factor DC4C1s

C1r

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macrophagesmicroglia

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