Constitutive Ret Activity in Knock-In Multiple Endocrine Neoplasia Type B Mice Induces Profound Elevation of Brain Dopamine Concentration via Enhanced Synthesis and Increases the Number of TH-Positive Cells in the Substantia Nigra

Abstract

Ret is the common signaling receptor for glial cell line-derived neurotrophic factor (GDNF) and other ligands of the GDNF family that have potent effects on brain dopaminergic neurons. The Met918Thr mutation leads to constitutive activity of Ret receptor tyrosine kinase, causing the cancer syndrome called multiple endocrine neoplasia type B (MEN2B). We used knock-in MEN2B mice with the Ret-MEN2B mutation to study the effects of constitutive Ret activity on the brain dopaminergic system and found robustly increased concentrations of dopamine (DA) and its metabolites in the striatum, cortex, and hypothalamus. The concentrations of brain serotonin were not affected and those of noradrenaline were slightly increased only in the lower brainstem. Tyrosine hydroxylase (TH) protein levels were increased in the striatum and substantia nigra/ventral tegmental area (SN/VTA), and TH mRNA levels were increased in SN/VTA of MEN2B mice, suggesting that constitutive Ret activity increases DA levels by increasing its synthesis. Also, the striatal DA transporter protein levels in the MEN2B mice were increased, which agrees with increased sensitivity of these mice to the stimulatory effects of cocaine. In the SN pars compacta of homozygous MEN2B mice, we found a 26% increase in the number of TH-positive cells, but no differences were found in the VTA. Thus, we show here that the constitutive Ret activity in mice is sufficient to increase the number of dopaminergic neurons and leads to profound elevation of brain DA concentration. These data clearly suggest that Ret activity per se can have a direct biological function that actively changes and shapes the brain dopaminergic system.

Pitx3 Is a Critical Mediator of GDNF-Induced BDNF Expression in Nigrostriatal Dopaminergic Neurons

“…Inactivation of the murine GDNF, Ret, and Gfra1 genes apparently does not interfere with prenatal mdDA neuron differentiation or survival (Airaksinen and Saarma, 2002;Kramer et al, 2007;Paratcha and Ledda, 2008). Moreover, persistent overexpression of GDNF driven by the TH promoter leads to a reduction of SNc DA neurons (Chun et al, 2002), whereas intrastriatal or intranigral injections of GDNF protein during early postnatal stages, or the constitutive activation of the Ret receptor, result in increased numbers of TH ϩ SNc neurons and an enhanced DA metabolism (Beck et al, 1996;Mijatovic et al, 2007). The transient but detectable expression of GDNF in the midgestational VM shown here might have been missed by previous in situ hybridization studies due to lower sensitivity or slight differences in the staging of the embryos (Hellmich et al, 1996;Golden et al, 1999), but GDNF expression in the embryonic rat VM was also shown by Choi-Lundberg and Bohn (1995) using a more sensitive RT-PCR-based detection method.…”

Death Receptors and Caspases But Not Mitochondria Are Activated in the GDNF- or BDNF-Deprived Dopaminergic Neurons

“…It is controversial whether GDNF actually controls the number of DA neurons during development (Janec and Burke, 1993;Jackson-Lewis et al, 2000;Krieglstein, 2004). Indeed, function-blocking antibodies to GDNF reduced the death of neonatal DA neurons in vivo (Oo et al, 2003(Oo et al, , 2005, but genetic manipulations of Ret affected adult rather than neonatal DA neurons (Granholm et al, 2000;Jain et al, 2006;Kramer et al, 2007;Mijatovic et al, 2007). More studies, including mice with conditional deletion of GDNF in the striatum, are required to solve the issue.…”

“…GDNF binds to coreceptor GFR␣1, and this complex activates receptor tyrosine kinase Ret (Bespalov and Saarma, 2007). Genetic manipulations of Ret in the DA neurons (Granholm et al, 2000;Jain et al, 2006;Kramer et al, 2007;Mijatovic et al, 2007) have given controversial results whether and/or when Ret physiologically regulates survival/death of DA neurons. Treatment of Parkinson's patients with GDNF has also been contradictory, because some studies reported considerable improvement (Gill et al, 2003;Slevin et al, 2005), whereas others did not (Lang et al, 2006).…”

Structure and Physiology of the RET Receptor Tyrosine Kinase

“…The robust effects of GDNF/RET signaling on dopaminergic neuron survival in several lesion paradigms naturally raised expectations of an important physiological function for RET in dopaminergic neurons, where it is expressed from very early stages of development. In agreement with this, knock-in of a constitutive allele of RET resulted in increased numbers of dopaminergic neurons and profound elevation of brain dopamine concentration, suggesting that RET signaling can have a direct biological effect in the brain dopaminergic system (Mijatovic et al 2007). Despite the successes of gain-offunction approaches, the results from loss-of-function studies have been less clear-cut with regard to the importance of RET activity in dopaminergic neurons.…”