Abstract
The growing field of RNA neurobiology dictates development and improvement of effective and reliable in vivo techniques to address the function of particular microRNA molecules within the brain. Here we describe a novel method involving continuous delivery of oligonucleotides into a brain region of interest by osmotic pump infusion. The approach implements application of double-stranded microRNA-mimics with only two LNA moieties at the 3′-end and additionally one at the 5′-end of the sense strand. This method holds promise for long-lasting and specific siRNA upregulation in vivo, especially in the Dicer-depleted systems, where other approaches are limited or not applicable. Being robust and effective, various techniques described in this chapter can be easily modified in order to achieve up- or downregulation of expression of specific RNA molecules, bi- or unilateral infusions or injections, and in vivo “screening” strategy allowing to start from a bigger group of RNA molecules and end up with identification of single RNA species critical for a phenotype.
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References
Bartel DP (2009) MicroRNAs: target recognition and regulatory functions. Cell 136(2):215–233. doi:10.1016/j.cell.2009.01.002, S0092-8674(09)00008-7 [pii]
Mikl M, Vendra G, Doyle M, Kiebler MA (2010) RNA localization in neurite morphogenesis and synaptic regulation: current evidence and novel approaches. J Comp Physiol A Neuroethol Sens Neural Behav Physiol 196(5):321–334. doi:10.1007/s00359-010-0520-x
Schratt G (2009) microRNAs at the synapse. Nat Rev Neurosci 10(12):842–849. doi:10.1038/nrn2763, nrn2763 [pii]
Konopka W, Schutz G, Kaczmarek L (2011) The microRNA contribution to learning and memory. Neuroscientist 17(5):468–474. doi:10.1177/1073858411411721
Konopka W, Kiryk A, Novak M, Herwerth M, Parkitna JR, Wawrzyniak M, Kowarsch A, Michaluk P, Dzwonek J, Arnsperger T, Wilczynski G, Merkenschlager M, Theis FJ, Kohr G, Kaczmarek L, Schutz G (2010) MicroRNA loss enhances learning and memory in mice. J Neurosci 30(44):14835–14842. doi:10.1523/JNEUROSCI.3030-10.2010, 30/44/14835 [pii]
Vinnikov IA, Hajdukiewicz K, Reymann J, Beneke J, Czajkowski R, Roth LC, Novak M, Roller A, Dörner N, Starkuviene V, Theis FJ, Erfle H, Schütz G, Grinevich V, Konopka W (2014) Hypothalamic miR-103 protects from hyperphagic obesity in mice. J Neurosci 34(32):10659–10674. doi:10.1523/jneurosci.4251-13.2014
Verma P, Augustine GJ, Ammar MR, Tashiro A, Cohen SM (2015) A neuroprotective role for microRNA miR-1000 mediated by limiting glutamate excitotoxicity. Nat Neurosci 18(3):379–385. doi:10.1038/nn.3935
Gao J, Wang WY, Mao YW, Graff J, Guan JS, Pan L, Mak G, Kim D, Su SC, Tsai LH (2010) A novel pathway regulates memory and plasticity via SIRT1 and miR-134. Nature 466(7310):1105–1109. doi:10.1038/nature09271, nature09271 [pii]
Tan CL, Plotkin JL, Veno MT, von Schimmelmann M, Feinberg P, Mann S, Handler A, Kjems J, Surmeier DJ, O’Carroll D, Greengard P, Schaefer A (2013) MicroRNA-128 governs neuronal excitability and motor behavior in mice. Science 342(6163):1254–1258. doi:10.1126/science.1244193
Xie J, Ameres SL, Friedline R, Hung JH, Zhang Y, Xie Q, Zhong L, Su Q, He R, Li M, Li H, Mu X, Zhang H, Broderick JA, Kim JK, Weng Z, Flotte TR, Zamore PD, Gao G (2012) Long-term, efficient inhibition of microRNA function in mice using rAAV vectors. Nat Methods 9(4):403–409. doi:10.1038/nmeth.1903
Ran FA, Cong L, Yan WX, Scott DA, Gootenberg JS, Kriz AJ, Zetsche B, Shalem O, Wu X, Makarova KS, Koonin EV, Sharp PA, Zhang F (2015) In vivo genome editing using Staphylococcus aureus Cas9. Nature. doi:10.1038/nature14299
Platt RJ, Chen S, Zhou Y, Yim MJ, Swiech L, Kempton HR, Dahlman JE, Parnas O, Eisenhaure TM, Jovanovic M, Graham DB, Jhunjhunwala S, Heidenreich M, Xavier RJ, Langer R, Anderson DG, Hacohen N, Regev A, Feng G, Sharp PA, Zhang F (2014) CRISPR-Cas9 knockin mice for genome editing and cancer modeling. Cell 159(2):440–455. doi:10.1016/j.cell.2014.09.014
Christensen M, Larsen LA, Kauppinen S, Schratt G (2010) Recombinant adeno-associated virus-mediated microRNA delivery into the postnatal mouse brain reveals a role for miR-134 in dendritogenesis in vivo. Front Neural Circuits 3:16. doi:10.3389/neuro.04.016.2009
Zovoilis A, Agbemenyah HY, Agis-Balboa RC, Stilling RM, Edbauer D, Rao P, Farinelli L, Delalle I, Schmitt A, Falkai P, Bahari-Javan S, Burkhardt S, Sananbenesi F, Fischer A (2011) microRNA-34c is a novel target to treat dementias. EMBO J 30(20):4299–4308. doi:10.1038/emboj.2011.327
Krützfeldt J, Rajewsky N, Braich R, Rajeev KG, Tuschl T, Manoharan M, Stoffel M (2005) Silencing of microRNAs in vivo with ‘antagomirs’. Nature 438(7068):685–689. doi:10.1038/nature04303
Krutzfeldt J, Kuwajima S, Braich R, Rajeev KG, Pena J, Tuschl T, Manoharan M, Stoffel M (2007) Specificity, duplex degradation and subcellular localization of antagomirs. Nucleic Acids Res 35(9):2885–2892. doi:10.1093/nar/gkm024, gkm024 [pii]
Esau C, Davis S, Murray SF, Yu XX, Pandey SK, Pear M, Watts L, Booten SL, Graham M, McKay R, Subramaniam A, Propp S, Lollo BA, Freier S, Bennett CF, Bhanot S, Monia BP (2006) miR-122 regulation of lipid metabolism revealed by in vivo antisense targeting. Cell Metab 3(2):87–98. doi:10.1016/j.cmet.2006.01.005
Liu J, Yu D, Aiba Y, Pendergraff H, Swayze EE, Lima WF, Hu J, Prakash TP, Corey DR (2013) ss-siRNAs allele selectively inhibit ataxin-3 expression: multiple mechanisms for an alternative gene silencing strategy. Nucleic Acids Res 41(20):9570–9583. doi:10.1093/nar/gkt693
Gatfield D, Le Martelot G, Vejnar CE, Gerlach D, Schaad O, Fleury-Olela F, Ruskeepaa AL, Oresic M, Esau CC, Zdobnov EM, Schibler U (2009) Integration of microRNA miR-122 in hepatic circadian gene expression. Genes Dev 23(11):1313–1326. doi:10.1101/gad.1781009
Rayner KJ, Esau CC, Hussain FN, McDaniel AL, Marshall SM, van Gils JM, Ray TD, Sheedy FJ, Goedeke L, Liu X, Khatsenko OG, Kaimal V, Lees CJ, Fernandez-Hernando C, Fisher EA, Temel RE, Moore KJ (2011) Inhibition of miR-33a/b in non-human primates raises plasma HDL and lowers VLDL triglycerides. Nature 478(7369):404–407. doi:10.1038/nature10486
Elmen J, Lindow M, Silahtaroglu A, Bak M, Christensen M, Lind-Thomsen A, Hedtjarn M, Hansen JB, Hansen HF, Straarup EM, McCullagh K, Kearney P, Kauppinen S (2007) Antagonism of microRNA-122 in mice by systemically administered LNA-antimiR leads to up-regulation of a large set of predicted target mRNAs in the liver. Nucleic Acids Res 36(4):1153–1162. doi:10.1093/nar/gkm1113
Elmén J, Lindow M, Schütz S, Lawrence M, Petri A, Obad S, Lindholm M, Hedtjärn M, Hansen HF, Berger U, Gullans S, Kearney P, Sarnow P, Straarup EM, Kauppinen S (2008) LNA-mediated microRNA silencing in non-human primates. Nature 452(7189):896–899. doi:10.1038/nature06783
Lanford RE, Hildebrandt-Eriksen ES, Petri A, Persson R, Lindow M, Munk ME, Kauppinen S, Orum H (2009) Therapeutic silencing of microRNA-122 in primates with chronic hepatitis C virus infection. Science 327(5962):198–201. doi:10.1126/science.1178178
Zhang Y, Wang Z, Gemeinhart RA (2013) Progress in microRNA delivery. J Control Release 172(3):962–974. doi:10.1016/j.jconrel.2013.09.015
Elmen J, Thonberg H, Ljungberg K, Frieden M, Westergaard M, Xu Y, Wahren B, Liang Z, Orum H, Koch T, Wahlestedt C (2005) Locked nucleic acid (LNA) mediated improvements in siRNA stability and functionality. Nucleic Acids Res 33(1):439–447. doi:10.1093/nar/gki193, 33/1/439 [pii]
Yu D, Pendergraff H, Liu J, Kordasiewicz HB, Cleveland DW, Swayze EE, Lima WF, Crooke ST, Prakash TP, Corey DR (2012) Single-stranded RNAs use RNAi to potently and allele-selectively inhibit mutant huntingtin expression. Cell 150(5):895–908. doi:10.1016/j.cell.2012.08.002
Mook OR, Baas F, de Wissel MB, Fluiter K (2007) Evaluation of locked nucleic acid-modified small interfering RNA in vitro and in vivo. Mol Cancer Ther 6(3):833–843. doi:10.1158/1535-7163.mct-06-0195
Mong JA, Devidze N, Goodwillie A, Pfaff DW (2003) Reduction of lipocalin-type prostaglandin D synthase in the preoptic area of female mice mimics estradiol effects on arousal and sex behavior. Proc Natl Acad Sci U S A 100(25):15206–15211. doi:10.1073/pnas.2436540100
Bramsen JB, Laursen MB, Damgaard CK, Lena SW, Ravindra Babu B, Wengel J, Kjems J (2007) Improved silencing properties using small internally segmented interfering RNAs. Nucleic Acids Res 35(17):5886–5897. doi:10.1093/nar/gkm548
Su J, Baigude H, McCarroll J, Rana TM (2011) Silencing microRNA by interfering nanoparticles in mice. Nucleic Acids Res 39(6):e38. doi:10.1093/nar/gkq1307
John M, Constien R, Akinc A, Goldberg M, Moon Y-A, Spranger M, Hadwiger P, Soutschek J, Vornlocher H-P, Manoharan M, Stoffel M, Langer R, Anderson DG, Horton JD, Koteliansky V, Bumcrot D (2007) Effective RNAi-mediated gene silencing without interruption of the endogenous microRNA pathway. Nature 449(7163):745–747. doi:10.1038/nature06179
Trajkovski M, Hausser J, Soutschek J, Bhat B, Akin A, Zavolan M, Heim MH, Stoffel M (2011) MicroRNAs 103 and 107 regulate insulin sensitivity. Nature 474(7353):649–653. doi:10.1038/nature10112, nature10112 [pii]
Lee ST, Chu K, Jung KH, Kim JH, Huh JY, Yoon H, Park DK, Lim JY, Kim JM, Jeon D, Ryu H, Lee SK, Kim M, Roh JK (2012) miR-206 regulates brain-derived neurotrophic factor in Alzheimer disease model. Ann Neurol 72(2):269–277. doi:10.1002/ana.23588
Cunningham MG, O’Connor RP, Wong SE (2008) Construction and implantation of a microinfusion system for sustained delivery of neuroactive agents. J Vis Exp. doi:10.3791/716, 716 [pii]
Zovoilis A, Agbemenyah HY, Agis-Balboa RC, Stilling RM, Edbauer D, Rao P, Farinelli L, Delalle I, Schmitt A, Falkai P, Bahari-Javan S, Burkhardt S, Sananbenesi F, Fischer A (2011) microRNA-34c is a novel target to treat dementias. EMBO J 30(20):4299–4308. doi:10.1038/emboj.2011.327, emboj2011327 [pii]
Paxinos G, Franklin KBJ (2001) The mouse brain in stereotaxic coordinates, 2nd edn. Academic Press, San Diego
Acknowledgments
The authors declare no competing financial interests. This work has been supported by the DFG through SFB488, the EU through grant LSHM-CT-2005-018652 (CRESCENDO), the BMBF through NGFNplus grants FZK-01GS08153 and 01GS08142, and the HGF through Initiative CoReNe (Network II, E2) the National Science Centre (Poland) grant (SONATA) 2011/01/D/NZ4/03744, grant (HARMONIA) 2013/08/M/NZ3/01045, and the Academy of Finland. We thank Jörg Krummheuer for the protocol for preparation of the LNA-oligonucleotides for injection as well as assistance in developing the oligonucleotide synthesis strategy, Günther Schütz for support, Lena Roth for assistance with the infusion technique. Special thanks goes to Adair Oesterle for assistance with techniques to fabricate micropipettes.
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Vinnikov, I.A., Domanskyi, A., Konopka, W. (2016). Continuous Delivery of Oligonucleotides into the Brain. In: Kye, M. (eds) MicroRNA Technologies. Neuromethods, vol 128. Humana Press, New York, NY. https://doi.org/10.1007/7657_2016_3
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DOI: https://doi.org/10.1007/7657_2016_3
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