1 Intrastriatal botulinum toxin abolishes pathologic rotational behaviour and induces 2 axonal varicosities in the 6-OHDA rat model of Parkinson's disease 3 Andreas Wree a,1 , Eilhard Mix b,1,2 , Alexander Hawlitschka a , Veronica Antipova a , Martin Witt a,c , 4 Oliver Schmitt a , Reiner Benecke b, ⁎ 5 a Department of Anatomy, University of Rostock, Germany 6 b Department of Neurology, University of Rostock, Gehlsheimer Str. 20, 18147 Rostock, Germany 7 c Department of Anatomy, University of Technology, Dresden, Germany 8 9 abstract article info 10 Article history: 11 Received 9 July 2010 12 Revised 14 September 2010 13 Accepted 23 September 2010 14 Available online xxxx 15 16 17 18 Keywords: 19 Botulinum neurotoxin A 20 6-Hydroxy-dopamine 21 Striatum 22 Motor function 23 Apomorphine-induced rotation 24 Axonal varicosities 25 Central pathophysiological pathways of basal ganglia dysfunction imply a disturbed interaction of 26 dopaminergic and cholinergic circuits. In Parkinson's disease (PD) imbalanced cholinergic hyperactivity 27 prevails in the striatum. Interruption of acetylcholine (ACh) release in the striatum by locally injected 28 botulinum neurotoxin A (BoNT-A) has been studied in the rat 6-hydroxydopamine (6-OHDA) model of PD 29 (hemi-PD). The hemi-PD was induced by injection of 6-OHDA into the right medial forebrain bundle. Motor 30 dysfunction provoked by apomorphine-induced anti-clockwise rotation was completely reversed for more 31 than 3 months by ipsilateral intrastriatal application of 1–2 ng BoNT-A. Interestingly, BoNT-A injected alone 32 into the right striatum of naïve rats caused a slight transient clockwise apomorphine-induced rotation, which 33 lasted only for about one month. Immunohistochemically, large axonal swellings appeared within the 34 striatum injected with BoNT-A, which we tentatively named BoNT-A-induced varicosities. They contained 35 either choline acetyltransferase or tyrosine hydroxylase. These findings suggest a selective inhibition of 36 evoked release of ACh by locally applied BoNT-A. Intrastriatal application of BoNT-A may antagonize localized 37 relative functional disinhibited hypercholinergic activity in neurodegenerative diseases such as PD avoiding 38 side effects of systemic anti-cholinergic treatment. 39 © 2010 Elsevier Inc. All rights reserved. 40 41 42 43 44 Introduction 45 A hallmark of Parkinson's disease (PD) is diminished dopaminergic 46 signaling in the striatum (caudate putamen, CPu), which leads to 47 increased release of acetylcholine (ACh) by disinhibited tonically 48 active interneurons with the consequence of a disturbed network 49 function and consecutive motor dysfunction. Current therapeutic 50 strategies are based on two major approaches in order to correct the 51 disturbed circuits of involuntary movement control, i.e. stimulation of 52 dopamine (DA) receptors on GABAergic medium spiny neurons and 53 inhibition of hypercholinergic activity of tonically active interneurons 54 (Day et al., 2006; Obeso et al., 2008). However, both approaches are 55 compromised by adverse side effects due to systemic drug application 56 (Whitney, 2007). In the pre-L-DOPA era, the first drugs that turned 57 out to have clinically significant anti-Parkinson activity were anti- 58 cholinergics such as atropine and atropine-like derivatives (Duvoisin, 59 1967; Kaplan et al., 1954; Ordenstein, 1868). Today some of these 60 compounds are still commonly used in clinical practice, e.g. the 61 piperidine derivative Biperiden (Akineton®). Also, the PD-like side 62 effects of anti-psychotics are typically treated with anti-cholinergic 63 drugs, especially in younger patients (Ekdawi and Fowke, 1966; 64 Mamo et al., 1999). However, the most serious problem with these 65 systemically administered anti-cholinergics remains the occurrence 66 of well-known troublesome peripheral and central side effects, such 67 as mydriasis, paresis of accommodation (blurred vision), reduced 68 salivation (dry mouth), parotitis, dry eyes, muscular pain, loss of 69 power, alteration of voice, dysphagia, regurgitation, constipation, 70 urinary retention, prostatism, tachycardia, fever in warm weather, 71 hallucinations, memory problems and confusion. To avoid this 72 disadvantage we tested local anti-cholinergic treatment applying 73 botulinum neurotoxin A (BoNT-A) into the CPu of hemiparkinsonian 74 (hemi-PD) rats, a well established 6-hydroxydopamine (6-OHDA)- 75 induced animal model of PD (Meredith et al., 2008). In this model, it 76 has been shown that intraperitoneally applied atropine antagonizes 77 the profound PD-typical akinesia (Schallert et al., 1978). In combina- 78 tion with L-DOPA it supported the alleviation of excessive bracing 79 reactions and suppressed pathological circling of the PD rats (Schallert 80 et al., 1979). In our study, intrastriatal application of BoNT-A caused 81 complete long-term abolition of pathological apomorphine-induced Neurobiology of Disease xxx (2010) xxx–xxx ⁎ Corresponding author. Fax: + 49 381 494 9512. Q2 E-mail addresses: eilhard.mix@med.uni-rostock.de (E. Mix), reiner.benecke@med.uni-rostock.de (R. Benecke). 1 These authors contributed equally to this work. 2 Fax: +49 381 494 9588. Available online on ScienceDirect (www.sciencedirect.com). YNBDI-02260; No. of pages: 8; 4C: 4, 5, 6 0969-9961/$ – see front matter © 2010 Elsevier Inc. All rights reserved. doi:10.1016/j.nbd.2010.09.017 Contents lists available at ScienceDirect Neurobiology of Disease journal homepage: www.elsevier.com/locate/ynbdi Please cite this article as: Wree, A., et al., Intrastriatal botulinum toxin abolishes pathologic rotational behaviour and induces axonal varicosities in the 6-HDA rat model of Parkinson's disease, Neurobiol. Dis. (2010), doi:10.1016/j.nbd.2010.09.017