Motor Dysfunction in Type 5 Adenylyl Cyclase-null Mice

• DOI: 10.1074/jbc.c300075200
• Publication Date: 2003-05-02T19:48:57Z
• Authors: Tamio Iwamoto; Satoshi Okumura; Kousaku Iwatsubo; Jun-Ichi Kawabe; Koji Ohtsu; Ikuko Sakai; Yoko Hashimoto; Aki Izumitani; Kazunori Sango; Kyoko Ajiki; Yoshiyuki Toya; Satoshi Umemura; Yoshio Goshima; Nobutaka Arai; Stephen F. Vatner; Yoshihiro Ishikawa
• Keywords: ADCY10; ADCY6; ADCY3

Various neurotransmitters, such as dopamine, stimulate adenylyl cyclase to produce cAMP, which regulates neuronal functions. Genetic disruption of the type 5 isoform led a major loss activity in striatum-specific manner with small increase expression few other isoforms. D1 dopaminergic agonist-stimulated was attenuated, and this accompanied by decrease receptor Gsα. D2 agonist-mediated inhibition also blunted. Type cyclase-null mice exhibited Parkinsonian-like motor dysfunction, i.e. abnormal coordination bradykinesia detected Rotarod pole test, respectively, lesser extent locomotor impairment open field tests. Selective or stimulation improved some these disorders mouse model, suggesting partial compensation each signal through remnant These findings extend our knowledge role an effector enzyme regulating signaling functions imply that provides site convergence both signals balances various wild cAMP-dependent protein kinase The neurotransmitter dopamine acts subtypes are associated either cyclases (ACs),1 leading regulation physiological control psychomotor (1Hardman J.G. Limbird L.E. Gilman A.G. Goodman ' Gilman's Pharmacological Basis Therapeutics. 10th Ed. McGraw-Hill, New York2001: 447-483Google Scholar). This dopamine-sensitive AC is highest striatum well limbic structures brain where their levels exceed, orders magnitude, those areas brain. Such differences striatal enzymatic may be attributed amount and/or combination isoforms expressed differentially region (2Chern Y. Cell. Signal. 2000; 12: 195-204Crossref PubMed Scopus (72) Google Scholar, 3Hanoune J. Defer N. Annu. Rev. Pharmacol. Toxicol. 2001; 41: 145-174Crossref (565) expresses all nine (AC1–AC9) have distinct biochemical properties,i.e. Gi, Gऔγ, calcium, kinases (4Taussig R. Zimmermann G. Adv. Second Messenger Phosphoprot. Res. 1998; 32: 81-98Crossref (47) Most, if not all, enriched specific regions 5Cooper D.M. Mons Karpen J.W. Nature. 1995; 374: 421-424Crossref (557) Scholar) rather than diffusely distributed throughout AC5, for example, dominant heart (6Premont R.T. Chen Ma H.W. Ponnapalli M. Iyengar Proc. Natl. Acad. Sci. U. S. A. 1992; 89: 9809-9813Crossref (170) 7Mons Decorte L. Jaffard Cooper Life 62: 1647-1652Crossref (76) 8Ishikawa Katsushika Halnon N.J. Kawabe Homcy C.J. Biol. Chem. 267: 13553-13557Abstract Full Text PDF However, coupling function remains unknown does whether isoform, unlike receptors, can substituted another isoform. considered center sensorimotor integration within basal ganglia (9Smith Kieval J.Z. Trends Neurosci. 23: S28-S33Abstract (251) receives widespread excitatory input from cortex converge extensive signals, D2, afferent midbrain. Concerted balanced two believed play key In study, we examined potential target use knockout AC5 gene disrupted. We disrupted homologous recombination technique at exon first translation initiation (Fig. 1A). has reasonable Kozak consensus sequence same (8Ishikawa excised final targeting vector. transgene confirmed genomic Southern analysis 1B). All were 129/SvJ-C57BL/6 mixed background littermates F1 heterozygote crosses. experiments performed 8–12-week-old homozygous (AC5−/−) wild-type (WT) littermates. study approved Animal Care Use Committee Yokohama City University School Medicine Jersey Medical School. Partial fragments cDNA clones (types 1–9) neuropeptides,i.e. enkephalin, substance P, dynorphin, obtained PCR. A human 28 S ribosomal RNA probe used internal control. RNase protection assay using RPA III kit (Ambion, Austin, TX). Striatal tissues dissected mice, membrane preparations prepared assays described previously (10Hess E.J. Battaglia Norman A.B. Creese I. Mol. 1987; 31: 50-57PubMed 11Ishikawa Sorota Kiuchi K. Shannon R.P. Komamura Vatner D.E. S.F. Clin. Investig. 1994; 93: 2224-2229Crossref (121) binding [3H]SCH23390 [3H]spiperone, (12Hess Iorio L.C. Eur. 1986; 121: 31-38Crossref (114) Scholar,13Baik J.H. Picetti Saiardi Thiriet Dierich Depaulis Le Meur Borrelli E. 377: 424-428Crossref (591) Preliminary demonstrated theKd Bmax values receptors similar reported 13Baik Motor assessed test (14Brandon E.P. Logue Adams M.R. Qi Sullivan S.P. Matsumoto A.M. Dorsa Wehner J.M. McKnight G.S. Idzerda R.L. 18: 3639-3649Crossref Scholar), tests 15Krezel W. Ghyselinck Samad T.A. Dupe V. Kastner P. Chambon Science. 279: 863-867Crossref (301) (16Matsuura Kabuto H. Makino Ogawa Methods. 1997; 73: 45-48Crossref (282) tail suspension (17Yamamoto Lucas J.J. Hen 101: 57-66Abstract (913) Given striatum, conducted evaluate animal model (AC5−/−). most dramatic change found coordinated movement, evaluated performance. measured time could stay on accelerating without falling. general, there AC5−/− degree heterozygous relative WT (Fig.2A). AC5−/−could spend significantly less Rotarod, slightly WT. When repeated, performance after trials. very poorly even several repeated following day, results similar, disputing possibility required longer period learn (data shown). Spontaneous determined horizontally (locomotion) vertically (rearings). Mice placed cage, movements videotaped analysis. revealed activity, while showed but significant reduction 2, B C). To bradykinesia, performed. until they turned downward (Tturn) descended floor (TLA). had marked deficits test; over 3-fold prolongation recording indexes (Fig.2D). It possible dysfunction choleric dystonic movements. readily clasping limbs triggered no indicated impairments presumably induced AC5. While regard its distribution (18Mons D.M.F. Brain 22: 236-244Crossref (95) it remained cAMP production striatum. mRNA least 10–20-fold more abundantly regions, cerebellum, (Fig.3A); agreement previous AC5−/−, negated, histological examinations changes reactive gliosis 8–12 weeks old found, however, greatly decreased AC5−/−(Fig. 3B). contrast, significantly, only degree, difference cerebellum scarcely For comparison, heart, tissue dominantly 307 shown), indicating contribution greater heart. (Fig.3C). many including attainable Gs-coupled agonists, although Gi-coupled agonists always easy. SKF38393, agonist, modestly stimulated (40.7 ± 2.67 10 ॖm GTP). Quinpirole, inhibited SKF38393-stimulated activity; (13.5 1.17 decrease). responses markedly diminished; small, hardly detectable. tentative speculate due Gi-inhibitable, proposed recently (19Lee K.W. Hong Choi I.Y. Che Lee J.K. Yang S.D. Song C.W. Kang H.S. Noh J.S. Shin Han P.L. 2002; 7931-7940Crossref catalytic too low demonstrate preparations. Thus, vitro sufficient terms sensitivity selective AC5−/−. did understand, why response agonist shown percent attenuated because must able respond Gs, stimulation. accumulation intact cells fetus; great above adults molecules involved signaling. sites (Fig.4A). short, long, form Gs 4B) positive feed forward regulatory loop. Western blotting molecules, preparation whole homogenates, Golf, Gq, Gऔ, PKA (the α subunit) changed Changes linked receptors. presynaptic κ-receptors inhibit (20Xie G.X. Meng F. Mansour Thompson R.C. Hoversten M.T. Goldstein Watson S.J. Akil 91: 3779-3783Crossref (70) increased 4C). expressions enkephalin P unchanged glutamic acid decarboxylase tyrosine hydroxylase, synthesis γ-aminobutyric immunoblotting suggested way suppress pathway despite then any Since antibodies convincingly determine level available, quantitated 4D). except AC4 AC8 detected. Among isoforms, modest AC6, relevant AC2, AC7, AC9 AC1 AC3 thought, increases explain decreases occurred pathway. If vivo, restore function. Administration SKF38393 (25 50 mg/kg), particular, appeared pronounced, might supersensitive, (Fig.5A). finding reminiscent supersensitive direct neurons observed depletion Parkinson's disease becomes actual (21Marshall J.F. Navarrete Joyce J.N. 1989; 493: 247-257Crossref (107) 22Gerfen C.R. Engber T.M. Mahan Susel Z. Chase T.N. Monsma Jr., F.J. Sibley D.R. 1990; 250: 1429-1432Crossref (2477) improve 5B). effect cabergoline (0.2 1.0 been treatment disease. Cabergoline tendency 5D). selectively AC5−/−; reached equivalent (Fig.5E), essentially WT, restored 5, C F). Both latter improvement lower dose mg/kg). deficit represented impaired mimicked coordination, attenuation underlied targets Locomotor stimulation, couple function, locomotion, respectively. Nevertheless disorders, essential balancing maintaining locomotion provide signals. abundant brain, output pathways,i.e. "direct" "indirect" pathways, mediated Although still populations (23Hersch S.M. Ciliax B.J. Gutekunst C.A. Rees H.D. Heilman Yung K.K. Bolam J.P. Ince Yi Levey A.I. 15: 5222-5237Crossref (24Surmeier D.J. Yan W.J. 42: 1020-1023Crossref (22) parallel activation pathways synergistic action Our indicate maintained presence coupled pathways. absence AC6 present cannot fully compensate (Fig.5A) supersensitivity As disease, Because up-regulation G expression, responsible supersensitization located downstream PKA, do deny include post-translational remaining components. exact molecular mechanisms paradoxical phenomenon unexplained recent switch mitogen-activated (25Gerfen Miyachi Paletzki Brown 5042-5054Crossref lack aspects, thus useful explore future studies. robust nature finding, suggests isoform,i.e. multiple subtype one equally important respective widely recognized. implicate isoform(s), pharmacotherapy effective treat diseases (26Onda T. Hashimoto Nagai Kuramochi Saito Yamazaki Toya Sakai Nishikawa Ishikawa 276: 47785-47793Abstract (102) thank Dr. Kazushige Touhara (University Tokyo) helpful discussion Hideaki Hori (Yokohama University) editorial assistance.