University of Zagreb School of Medicine

Laboratory of Molecular Neuropharmacology, Department of Pharmacology, University of Zagreb School of Medicine

PI:

Prof.dr.sc. Zdravko Lacković, M.D. (Department of Pharmacology, University of Zagreb School of Medicine, Zagreb, Croatia)

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Project team:

Prof.dr.sc. Maja Relja, M.D. (Department of Neurology, Clinical Hospital Center, University of Zagreb School of Medicine, Zagreb, Croatia)

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Prof.dr.sc. Lidija Bach-Rojecky, MPharm. (Department of Pharmacology, University of Zagreb School of Pharmacy and Biochemistry, Zagreb, Croatia)

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Dr.sc. Ivica Matak, dipl.ing.biol. (Department of Pharmacology, University of Zagreb School of Medicine, Zagreb, Croatia)

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Dr.sc. Boris Filipović, M.D. (Department of Pharmacology, University of Zagreb School of Medicine, Zagreb, Croatia)

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Dr.sc. Višnja Drinovac-Vlah, MPharm. (Department of Pharmacology, University of Zagreb School of Pharmacy and Biochemistry, Zagreb, Croatia)

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Dr.sc. Nataša Klepac, M.D. (Department of Neurology, Clinical Hospital Center, University of Zagreb School of Medicine, Zagreb, Croatia)

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Technician:

Božica Hržan  (Department of Pharmacology, University of Zagreb School of Medicine, Zagreb, Croatia)

Foreign collaborators:

Prof.dr.sc. Zsuzsanna Helyes, M.D. (Department of Pharmacology and Pharmacotherapy, Medical School, University of Pécs, Pécs, Hungary).

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Prof.dr.sc. Claudia Sommer, M.D. (Department of Neurology, University of Wurzburg, Würzburg, Germany)

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Dr.sc. Matteo Caleo, MSc (CNR Institute of Neuroscience, Pisa, Italy)

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Botulinum toxins (BTX) and tetanus toxin (TTX) are the most important clostridial neurotoxins and one of the most toxic substances in nature. One gram of BTX type A (BTX-A) could kill more than one million people via inhalational route. At the same time it is often used for therapeutic, especially cosmetic purposes (Botox).

Today, BTX-A use in certain types of pain is increasing. Long-term efficacy and safety are of a special clinical value. Until recently it was believed that the primary mechanism of BTX-A antinociceptive action is localized at peripheral nerve endings, mediated by prevention of neurotransmitters release, caused by cleavage of SNAP-25, protein required for exocytosys. Our laboratory revealed for the first time that BTX-A is retrogradely transported from periphery through sensory nerves to the CNS and that the site of its action on pain is central. However, the exact mechanism is yet unresolved. Thus, our project proposal consists of 7 mutually connected objectives:

1. Our recently published data suggested that opioid and GABA-ergic systems are involved in BTX-A central antinociceptive activity, via an unknown mechanism. Therefore, we will investigate: BTX-A influence on the expression of opioid receptors and opioid peptides precursor molecules; possible involvement of glutamate receptors in central BTX-A antinociceptive action; potential role of spinal and supraspinal glia in antinociceptive action of BTX-A;
2. We previously reported BTX-A antinociceptive effect on bilateral pain models. Our goals in the present project will be to investigate the mechanism of BTX-A action on bilateral pain and its possible interference with the central sensitization.

III. Our preliminary data also suggested that capsaicin-sensitive primary afferents are involved in central BTX-A antinociceptive activity. Therefore, we will further investigate association of capsaicin-sensitive neurons and BTX-A antinociceptive activity.

1. We recently discovered that different types of trigeminal experimental pain are accompanied by neurogenic inflammation of dura mater – the presumed primary mechanism of migraine headache. Herein we will investigate possible effect of BTX-A on trigeminovascular system and calcitonin gene-related peptide, as a possible mechanism of its antimigraine action.
2. In addition to sensory system, the central effects of BTX-A in motor system will be investigated too, which might lead to the better understanding of its beneficial effects on movement disorders.
3. Additionally, another clostridial toxin will be explored: the TTX. Although it has clinically opposite effects (spastic vs. flaccid paralysis), its molecular effects at the level of synapses are very similar to BTX. We assume that the selective delivery of small amounts of TTX into sensory nerves and ganglia could also have the effect on pain.

VII. In parallel, the effect of BTX-A on various non-motor functions in migraine or dystonic patients will be studied. This makes our research also a translational research.

Proposed research project could lead to the explanation of BTX-A mechanisms of action in chronic pain, migraine, and hyperkinetic movement disorders, which would make an important contribution to possible refinement of its clinical use. As a problem-oriented research, it requires various procedures and techniques (animal behavior, Western blot, immunohistochemistry, transgenics, PCR, etc.)

List of scientific publications resulting from the project:

2016-2017

Matak I, Tékus V, Bölcskei K, Lacković Z, Helyes Z. Involvement of substance P in the antinociceptive effect of botulinum toxin A. Neuroscience; 358(2017):137-145. Open Access

Link

Drinovac Vlah V, Filipović B, Bach-Rojecky L, Lacković Z. Role of central versus peripheral opioid system in antinociceptive and anti-inflammatory effect of botulinum toxin type A in trigeminal region.

Eur J Pain. 2017 Nov 13. doi: 10.1002/ejp.1146. (Article in press)

Drinovac Vlah et al. 2017 EJP

Relja M, Miletić V. When movement disorders hurt: Addressing pain in hyperkinetic disorders. Parkinsonism and related disorders; doi: 10.1016/j.parkreldis.2017.08.011.; 2017

Relja and Miletic 2017. Parkinsonism Relat Disord

2015-2016

Matak I, Lacković Z, Relja M. Botulinum toxin type A in motor nervous system: unexplained observations and new challenges. J Neural Transm (Vienna). 2016 Dec;123(12):1415-1421.

Matak et al. 2016 JNT

Drinovac Vlah V, Bach-Rojecky L, Lacković Z. (2016) Antinociceptive action of botulinum toxin type A in carrageenan-induced mirror pain. J Neural Transm (Vienna). 123(12):1403-1413

Drinovac Vlah et al. 2016 JNT

Congress abstracts:

2016-2017

Lacković Z,  Matak I, Filipović B. Pain induced inflammation of the meninges.  American Academy of Neurology Meeting 2017, 22-28.04.2017. Boston, USA

Lackovic et al. 2017 AAN

Bach-Rojecky L, Drinovac Vlah V, Lacković Z. Botulinum toxin type A and NMDA antagonist: additive antinociceptive effect at the spinal level. European Pain Federation Meeting EFIC 2017, 6.-9.09.2017. Copenhagen, Denmark

Bach-Rojecky et al. 2017 EFIC

Matak I, Drinovac-Vlah V, Lacković Z. Behavioral investigation of the effect of tetanus neurotoxin on sensory transmisssion in the rat facial region. FENS Regional Meeting, 20-23.9.2017. Pécs, Hungary

Link

2015-2016

Drinovac V, Bach-Rojecky L, Lacković Z. Antinociceptive effect of botulinum toxin type A on persistent bilateral allodynia induced by intramuscular carrageenan in rats. 8 th Croatian Congress of Pharmacology, 15-18.09.2016. Split, Croatia

Drinovac Vlah et al. 2016 Split

Lacković Z,  Bach-Rojecky L, Matak I, Helyes Z, Filipović B, Drinovac V, Relja M. Central mechanism of botulinum toxin action on pain supersensitivity, allodynia and migraine. European Congress of Pharmacology EPHAR 24.-28-06. 2016. Istanbul, Turkey

Lackovic et al. 2016 EPHAR

Relja M, Maravić M, Jurjević I, Matak I Optimising the effect of botulinum toxin treatment by post injection activity. European Congress of Pharmacology EPHAR 24.-28-06. 2016. Istanbul, Turkey

Relja et al. 2016 EPHAR

Lacković Z, Filipović B, Matak I, Helyes Z. Unexpected meningeal reaction to pain in trigeminal region: association with calcitonin gene related polipeptide. World Congress of Neurology 31.10 – 5.11.2015. Santiago de Chile, Chile

Link

Doctoral theses:

Boris Filipović (2016) Neurogena upala moždanih ovojnica i bol u području glave i vrata [Neurogenic inflammation of cranial meninges and pain in craniocervical area]. Mentor: prof.  Zdravko Lacković

Višnja Drinovac-Vlah (2017) Središnji neurotransmitori i mehanizam antinociceptivnog djelovanja botulinum toksina tipa A [Central neurotransmitters and the mechanisms of antinociceptive activity of botulinum toxin type A]. Mentors: prof. Zdravko Lacković and prof. Lidija Bach-Rojecky.

Link

New collaborations:

As a result of BrainTox project,  and previous work of our Laboratory,  a collaboration on botulinum toxin activity was started (signed August 1, 2017) with  pharmaceutical company Ipsen (Ipsen Innovation SAS, 5 Avenue du Canada, Zone industrielle de Courtaboeuf, Les Ulis, 91940, France, Ipsen Ref. Number 47504).