Fibroblast growth factor 20 is protective towards dopaminergic neurons in vivo in a paracrine manner

Abstract

Neuroprotective strategies are an unmet medical need for Parkinson's disease. Fibroblast growth factor 20 (FGF20) enhances survival of cultured dopaminergic neurons but little is known about its in vivo potential. We set out to examine whether manipulation of the FGF20 system affected nigrostriatal tract integrity in rats, to identify which fibroblast growth factor receptors (FGFRs) might reside on dopaminergic neurons and to discover the source of endogenous FGF20 in the substantia nigra (SN). Male Sprague Dawley rats were subject to a partial 6-OHDA lesion alongside treatment with exogenous FGF20 or an FGFR antagonist. Behavioural readouts and tyrosine-hydroxylase (TH) immunohistochemistry were used to evaluate nigrostriatal tract integrity. Fluorescent immunohistochemistry was used to examine FGFR subtype expression on TH-positive dopamine neurons and FGF20 cellular localisation within the SN. FGF20 (2.5 μg/day) significantly protected TH-positive cells in the SN and terminals in the striatum, while reducing the development of motor asymmetry at 5, 8 and 11 days post lesion. Conversely, the FGFR antagonist PD173074 (2 mg/kg) significantly worsened both the 6-OHDA lesion and resultant motor asymmetry. Within the SN, TH-positive cells expressed FGFR1, 3 and 4 while FGF20 co-localised with GFAP-positive astrocytes. In conclusion, FGF20 protects dopaminergic neurons in vivo, an action likely mediated through activation of FGFRs1, 3 or 4 found on these neurons. Given FGF20 is localised to astrocytes in the adult SN, endogenous FGF20 provides its protection of dopamine neurons through a paracrine action. Boosting the endogenous FGF20 production might offer potential as a future therapeutic strategy in Parkinson's disease.

Keywords: 6-Hydroxydopamine; Fibroblast growth factor 20; Fibroblast growth factor receptor; Neuroprotection; Parkinson's disease.

Fig. 1

FGF20 protects against dopamine neuron loss and motor deficits in the partial lesion 6-OHDA rat model. Quantification of (A) TH-positive cells in the lesioned SNc and (B) TH immunoreactivity in the lesioned striatum, both expressed as a percentage of the intact hemisphere. (C) Representative images of TH staining in the SNc and striatum. Arrow heads indicate the position of the SNc in the lesioned hemispheres. (D) Quantification of the behavioural asymmetry as measured using cylinder reaching. Data are mean ± SEM, n = 10 (vehicle) or 6 (FGF20 treatment groups); *P < 0.05 compared to vehicle-treated group (A and B); ***P < 0.001 versus pre-lesion score (day -1).

Fig. 2

FGFR antagonist, PD173074, exacerbates a partial 6-OHDA lesion in rats. Quantification of (A) TH-positive cells in the lesioned SNc and (B) TH immunoreactivity in the lesioned striatum, both expressed as a percentage of the intact hemisphere. (C) Quantification of the total number of amphetamine-induced rotations. Data are mean ± SEM, n = 7 (vehicle) or 6 (PD173074 treatment groups); *P < 0.05 compared to vehicle-treated group (A and C).

Fig. 3

FGF20 co-localises with astrocytes, but not dopamine neurons, in the SN. Immunofluorescent images showing (A) lack of FGF20 (red) signal on TH-positive neurons (green) in the SNc and (B) co-localisation of FGF20 (red) with GFAP (green) in the neighbouring SNr. Images were acquired at:×5 magnification (right hand panel in A; scale bar = 200 μm);×20 magnification (left panel in A; scale bar = 50 μm);×40 magnification (B; scale bar = 20 μm). (For interpretation of the references to colour in this figure legend, the reader is referred to the Web version of this article.)

Fig. 4

FGFR1, 3 and 4 are expressed on dopaminergic neurons in the SNc. Immunofluorescent images showing co-localisation of (A) FGFR1, (B) FGFR2, (C) FGFR3 and (D) FGFR4 with TH in coronal sections of the adult rat SNc counterstained with the nuclear marker, Hoechst (blue). Intense staining of FGFR1 (red), FGFR3 (green) and FGFR4 (green) is notable on TH-positive stained cells, while FGFR2 (red) shows no co-localisation with TH. All images were acquired at ×40 magnification. Scale bar = 20 μm. (For interpretation of the references to colour in this figure legend, the reader is referred to the Web version of this article.)