Apomorphine‐induced activation of dopamine receptors modulates FGF‐2 expression in astrocytic cultures and promotes survival of dopaminergic neurons

Abstract

Apomorphine (APO), a potent D1/D2 dopamine receptor agonist, is currently used as an antiparkinsonian drug. We have shown previously that APO stimulates synthesis and release of multiple trophic factors, such as brain-derived neurotrophic factor (BDNF) and glial cell line-derived neurotrophic factor (GDNF), in both mesencephalic and striatal neurons, thereby effectively preventing dopaminergic neuron loss in vitro. The present study was designed to investigate the effects of APO on fibroblast growth factor-2 (FGF-2) expression and regulation in astrocytes, and furthermore, to identify signaling mechanisms underlying these effects. Here, we show that FGF-2 expression is robustly induced in cultured astrocytes in response to APO. FGF-2 expression was proportional to APO concentration and time-dependent. Conversely, treatment with S-APO, a derivative of R-APO lacking DA receptor agonist activity, did not alter FGF-2 levels. APO treatment resulted in enhanced cytosol FGF-2 immunoreactivity, export of high MW forms of FGF-2 to the cytoplasm from the nucleus and increased extracellular release of FGF-2. Interestingly, both high and low MW forms of FGF-2 were detectable in conditioned medium of APO-treated cultures. This APO-induced effect was correlated with activation of D1 and D2 receptors, as it could be either mimicked by dopamine receptor agonists (SKF38393, quinpirole) or partially blocked by antagonists (SCH23390, SKF83566, haloperidol). Activation of the D1 receptor preferentially increased PKA activity, whereas activation of the D2 receptor only promoted phosphorylation of MAPK. Importantly, APO-modulated FGF-2 expression was independent of Akt/phosphoinositide 3-kinase signaling. These data suggest that APO can enhance biosynthesis and release of FGF-2 through activation of dopamine receptors in striatal astrocytes. Both cAMP/PKA and MEK/MAPK signaling cascades are major steps mediating this process.

Activation of Phosphatidylinositol-Linked D1-Like Receptor Modulates FGF-2 Expression in Astrocytes via IP3-Dependent Ca2+ Signaling

“…The conditioned medium (CM) was prepared as described previously (Li et al, 2006). Postnatal striatal astrocytes were allowed to grow to 90% confluence and maintained with FBS-free DMEM/Ham's F12 medium containing SKF83959 or APO for 24 h after pretreatment with or without 50 M SCH23390.…”

“…Western blotting was performed as described previously (Li et al, 2006). The following primary antibodies were used: (a) rabbit anti-FGF-2 pAb (1:300; Santa Cruz Biotechnology); (b) rabbit anti-BDNF pAb (1:200; Santa Cruz Biotechnology); (c) rabbit anti-GDNF pAb (1:500; Santa Cruz Biotechnology); (d) rabbit anti-neurotrophin-3 (NT-3; 1:200; Santa Cruz Biotechnology); (e) mouse anti-␣-tubulin monoclonal antibody (mAb; 1:4000; Sigma); (f) mouse anti-␤-actin mAb (1:5000; Sigma); (g) rabbit anti-phosphoCaMKII␣ pAb (1:500; Cell Signaling Technology).…”

“…Furthermore, APO was found to enhance biosynthesis and release of fibroblast growth factor-2 (FGF-2) which promotes the survival and protection from neurotoxin-induced death of DA neurons (Chadi et al, 1993;Grothe et al, 2000). Enhanced FGF-2 expression is further demonstrated to be mediated through activation of DA receptors in striatal astrocytes (Li et al, 2006). Together, these findings suggest a crucial role of nigrostriatal astrocytes in DA neuronal survival and the nigrostriatal astrocytes emerge as a promising target for drug development of PD.…”

Driving GDNF expression: The green and the red traffic lights

“…Besides the increase in GDNF and BDNF levels, treatment with apomorphine also increases FGF-2 expression in striatal astrocytes, both at the mRNA and protein level, an effect correlated with the activation of D1 and D2 receptors (Li et al, 2006a). FGF-2 exerts a potent neuroprotective action on nigral dopaminergic neurons in the MPTP mouse model of PD Unsicker, 1990, 1993;Chadi et al, 1993).…”

Unilateral 6-OHDA <i>th-fgfr1</i>(<i>tk-</i>) mouse model supports the role of FGFs in Parkinson’s disease and the effects of nicotine and L-DOPA on spontaneous motor impairments

“…FGF distinguishes itself from other neurotrophic factors due to its critical role in supporting developing and adult DA systems [22,23]. FGF-2 promotes the growth and survival of cultured DA neurons [24][25][26], protects against neurotoxin-induced cell death [27][28][29][30][31][32] and cografting of DA neurons with FGF-2 increases neuronal survival [17,33,34]. It was reported that FGF signaling increases DA levels in calbindin-negative SN neurons, those at greatest risk in PD patients [35].…”