Introduction that nicotine could have a positive effect in several

Brain Research 888 2001 336–342 www.elsevier.com locate bres Interactive report Nicotine prevents striatal dopamine loss produced by 1 6-hydroxydopamine lesion in the substantia nigra Gustavo Costa, J.A. Abin-Carriquiry, Federico Dajas ´ ´ ´ Division Neuroquımica , Instituto de Investigaciones Biologicas Clemente Estable, Avda. Italia 3318, 11 600 Montevideo, Uruguay Accepted 23 October 2000 Abstract While the work of several groups has shown the neuroprotective effects of nicotine in vitro, evidences for the same effects in vivo are controversial, mainly regarding neuroprotection in experimental models of Parkinson’s disease. In this context, we investigated the capability of various systemic administration schedules of nicotine to prevent the loss of striatal dopamine levels produced by partial or extensive 6-hydroxydopamine 6-OHDA lesion of rat substantia nigra SN. Eight days after 6- and 10-mg injections of 6-OHDA in the SN there was a significant decrease of dopamine concentrations in the corpus striatum CS and a concomitant increase in dopamine turnover. While 10 mg 6-OHDA produced an almost complete depletion of dopamine in the SN, 6 mg decreased dopamine levels by 50. Subcutaneous nicotine 1 mg kg administered 4 h before and 20, 44 and 68 h after 6 mg 6-OHDA, prevented significantly the striatal dopamine loss. Administered only 18 or 4 h before or only 20, 44 and 68 h after, nicotine failed to counteract the loss of dopamine or the increase in dopamine turnover observed in the CS. Nicotine also failed to prevent significantly the decrease of striatal dopamine levels produced by the 10-mg 6-OHDA intranigral dose. Chlorisondamine, a long-lasting nicotinic acetylcholine receptor antagonist, reverted significantly the nicotinic protective effects on dopamine concentrations. These results are showing that putative neuroprotective effects of nicotine in vivo depend on an acute intermittent administration schedule and on the extent of the brain lesion.  2001 Elsevier Science B.V. All rights reserved. Theme : Disorders of the nervous system Topic : Degenerative disease: Parkinson’s Keywords : Nicotine; Dopamine; Corpus striatum; Substantia nigra; Nicotinic acetylcholine receptor; 6-Hydroxydopamine; Chlorisondamine

1. Introduction that nicotine could have a positive effect in several

neuronal processes like attention, memory processing, Although initially introduced in Europe with tobacco pain, or neuroprotection [9]. The identification of more plants for medicinal purposes, nicotine has become, than 10 genes codifying the different sub-units of the through tobacco addiction, a harmful presence in our nAChR gave a structural basis for the search of specific modern societies. The physiological effects of nicotine are subunit combinations with beneficial effects in the CNS varied, affecting the peripheral cardiovascular, neuro- [9]. muscular and the central nervous system CNS [18], and Several reports have shown that there is a negative are based on its ability to mimic the actions of acetyl- correlation between smoking and the appearance of Parkin- choline at the nicotinic acetylcholine receptors nAChR. son’s disease. These epidemiological findings gave support The deleterious effects of tobacco use have precluded a to the hypothesis of a neuroprotective role of nicotine comprehensive study of the effects of nicotine in the CNS. [12,36] and prompted the study of the effects of nicotine In recent decades, however, several studies demonstrated stimulation in several models of neurotoxicity. At present, numerous studies have confirmed the protection conferred 1 by nicotine to neuronal cultures against toxic insults like Published on the World Wide Web on 1 December 2000. excitotoxins [1,11,22,31,35,42] or b-amyloid toxicity [29]. Corresponding author. Tel. fax: 1598-2-487-2603. E-mail address : fdajasiibce.edu.uy F. Dajas. Blockade of the neuroprotection effects by nAChR antago- 0006-8993 01 – see front matter  2001 Elsevier Science B.V. All rights reserved. P I I : S 0 0 0 6 - 8 9 9 3 0 0 0 3 0 8 7 - 0 G . Costa et al. Brain Research 888 2001 336 –342 337 nists provided evidence for the specific mediation by in aCSF containing 0.2 ascorbic acid. Nicotine and CHL different subtypes of nAChR [29,31,43]. were dissolved in saline. In spite of this in vitro evidence, in vivo studies are still controversial when regarding the neuroprotective prop- 2.3. Intranigral injection of 6-OHDA erties of nicotine treatments. While continuous nicotine infusion has been demonstrated to protect against neuronal Animals were anaesthetized with halothane Fluothane, loss produced by dopamine pathways hemitransection Zeneca and placed in a D. Kopf stereotaxic frame. [25,26], the striatal depletion of dopamine after injection of Through a skull hole, the needle 0.022 mm o.d., 0.013 6-hydroxydopamine 6-OHDA in the substantia nigra mm i.d. of a Hamilton syringe 5 ml, attached to a SN was unaltered by the same treatment [10]. 1-Methyl- micro-injection unit D. Kopf, was lowered to the SN. 4-phenyl-1,2,3,6-tetrahydropyridine MPTP systemic ap- Coordinates H, 24.8; L, 22.2; V, 27.2 were determined plication in vivo resulted in a significant decrease of from bregma, according to the atlas of Paxinos and Watson striatal dopamine content that was not prevented by [37]. nicotine in some studies [21], while another group showed A total of 2.0 ml of a 6-OHDA solution 3 and 5 mg ml that nicotine had protective effects against diethyldithio- for the 6- and 10-mg doses was injected for 4 min and the carbamate enhancement of MPTP lesions [34]. Moreover, needle was slowly withdrawn, allowing the drug to diffuse in the MPTP model of experimental parkinsonism, nicotine for another minute. Body temperature was maintained at has even shown an enhancement of the neurotoxicity 378C using a temperature control system. [5,19]. Beyond these discrepancies, it is likely that the great 2.4. Experimental groups and nicotine administration variety of experimental conditions reported, differing in schedule the schedule and method of nicotine administration, may in part explain the differences observed. Thus, reports using 2.4.1. Partial lesion 6 mg 6-OHDA in the SN chronic treatments did not show effects in vivo [21,27], Five groups of rats n58 injected with 6-OHDA 6 mg while the acute intermittent administration appeared to in the right SN, received 1 mg kg nicotine subcutaneously show protective effects in the models reported [27,34]. according to the following protocols: 1 nicotine 18 h In this context, and as a contribution to the characteriza- before 6-OHDA; 2 nicotine 4 h before 6-OHDA; 3 tion of the role played by nAChR in Parkinson’s disease, nicotine 4 h before, and 20, 44 and 68 h after 6-OHDA; 4 we studied the putative neuroprotective effects of nicotine nicotine 20, 44, 68 h after 6-OHDA; 5 similar adminis- in the 6-OHDA model of experimental parkinsonism, tration schedule to 3 plus CHL 10 mg kg s.c. 30 min assessing whether the timing and schedule of nicotine before the first application of nicotine. administration as well as the extent of the lesion are factors Each of the five experimental groups had its own control that could effectively influence the neuroprotection profile. group of 6-OHDA plus saline instead of nicotine. For control of 6-OHDA vehicle, six rats were injected with similar volumes of aCFS with 0.2 ascorbic acid.

2. Materials and methods