Neuroinflammation mediated by IL-1beta increases susceptibility of dopamine neurons to degeneration in an animal model of Parkinson's disease

Abstract

Background: The etiology of Parkinson's disease (PD) remains elusive despite identification of several genetic mutations. It is more likely that multiple factors converge to give rise to PD than any single cause. Here we report that inflammation can trigger degeneration of dopamine (DA) neurons in an animal model of Parkinson's disease.

Methods: We examined the effects of inflammation on the progressive 6-OHDA rat model of Parkinson's disease using immunohistochemistry, multiplex ELISA, and cell counting stereology.

Results: We show that a non-toxic dose of lipopolysaccharide (LPS) induced secretion of cytokines and predisposed DA neurons to be more vulnerable to a subsequent low dose of 6-hydroxydopamine. Alterations in cytokines, prominently an increase in interleukin-1beta (IL-1beta), were identified as being potential mediators of this effect that was associated with activation of microglia. Administration of an interleukin-1 receptor antagonist resulted in significant reductions in tumor necrosis factor-alpha and interferon-gamma and attenuated the augmented loss of DA neurons caused by the LPS-induced sensitization to dopaminergic degeneration.

Conclusion: These data provide insight into the etiology of PD and support a role for inflammation as a risk factor for the development of neurodegenerative disease.

Figure 1

Experimental timelines. Each timeline represents in vivo experimental procedures and postmortem analyses conducted. Number of animals in each group: (A) n = 6, (B) n = 6, (C) n = 7, (D) n = 7, (E) n = 5, (F) n = 8. LPS, lipopolysaccharide; 6-OHDA, 6-hydroxydopamine, SAL, saline; IL-1ra, interluekin-1 receptor antagonist; veh, vehicle; SN, substantia nigra, Str, striatum.

Figure 2

(A) Tyrosine hydroxylase positive (TH+) cell counts in the substantia nigra (SN). (B) Animals received, into the SN, saline (SAL) and 12 days later received an intra-striatal injection of SAL. (C) Animals received lipopolysaccharide (LPS) into the SN and 12 days later received an intra-striatal injection of SAL. (D) Animals received SAL into the SN and 12 days later received an intra-striatal injection of 6-OHDA (5.0 μg). (E) Animals received LPS into the SN and 12 days later received an intra-striatal injection of 6-OHDA (5.0 μg). (F) Animals received an intra-striatal injection of 6-OHDA (5.0 μg) and 12 days later received LPS into the SN. (G) Animals received an intra-striatal injection of 6-OHDA-h (high) (22.5 μg). Letters inside bars of panel A represent respective images in panels below. 21 days was allowed following 6-OHDA injection in all conditions. There was an overall main effect across groups (ANOVA, p < 0.001). LPS injection into the SN was non-toxic to TH+ neurons (B vs. C, NS). LPS injection prior to 6-OHDA administration increased the amount of TH+ cell loss compared to SAL prior to 6-OHDA (D vs. E, *p < 0.05, post-hoc Holm-Sidak). Intra-striatal injection of 6-OHDA followed by injection of LPS in the SN produced no greater cell loss (D vs. F, NS). The high dose of 6-OHDA produced cell loss that was not significantly different from a low dose of 6-OHDA with prior exposure to LPS (E vs. G, NS), however the high dose of 6-OHDA was significantly greater than other conditions receiving the lower dose of 6-OHDA (G vs. D and F, # p < 0.05, post-hoc Holm-Sidak). 1^st, first injection; 2^nd, second injection; SNpc, pars compacta; SNpr, pars reticulata; VTA, ventral tegmental area; CP, cerebral peduncle; scale bar, 1.0 mm; magnification, 2.5×. Error bars, ± SEM; n = 6 per condition.

Figure 3

Representative coronal sections of microglia in the ventral midbrain at the level of the substantia nigra (SN) visualized by CD11b-immunoreactivity (-ir). (A) activated microglia in the SN of a rat injected in the SN with LPS 12 days prior and (B) the contralateral side. (C) low level of microglia activation 12 days following an intra-striatal injection of 6-OHDA (5.0 μg) and (D) the contralateral side showing resting microglia. Insets are 20× images taken from the area outlined in lower magnification pictures. Scale bar of inset, 100 μm; scale bar of low magnification images (2.5×), 1.0 mm).

Figure 4

Interleukin-1 receptor antagonist (IL-1ra) reduces the amount of tyrosine hydroxylase-immunoreactive (TH-ir) cell loss associated with LPS and 6-OHDA (t-test, *p < 0.05). Each animal received LPS into the substantia nigra (SN), 9 days later started on either IL-1ra or vehicle (Veh) which continued through the experiment, and all animals then received an intra-striatal injection of 6-OHDA (5.0 μg) and were allowed 21 days until post-mortem analyses. Magnification, 2.5×; scale bar, 1.0 mm. Error bars, ± SEM; n = 8 per condition.