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. 2013 Nov 20;3:3273.
doi: 10.1038/srep03273.

Oral 'Hydrogen Water' Induces Neuroprotective Ghrelin Secretion in Mice

Free PMC article

Oral 'Hydrogen Water' Induces Neuroprotective Ghrelin Secretion in Mice

Akio Matsumoto et al. Sci Rep. .
Free PMC article


The therapeutic potential of molecular hydrogen (H₂) is emerging in a number of human diseases and in their animal models, including in particular Parkinson's disease (PD). H₂ supplementation of drinking water has been shown to exert disease-modifying effects in PD patients and neuroprotective effects in experimental PD model mice. However, H₂ supplementation does not result in detectable changes in striatal H₂ levels, indicating an indirect effect. Here we show that H₂ supplementation increases gastric expression of mRNA encoding ghrelin, a growth hormone secretagogue, and ghrelin secretion, which are antagonized by the β1-adrenoceptor blocker, atenolol. Strikingly, the neuroprotective effect of H₂ water was abolished by either administration of the ghrelin receptor-antagonist, D-Lys(3) GHRP-6, or atenolol. Thus, the neuroprotective effect of H₂ in PD is mediated by enhanced production of ghrelin. Our findings point to potential, novel strategies for ameliorating pathophysiology in which a protective effect of H₂ supplementation has been demonstrated.


Figure 1
Figure 1. Oral H2 water increases ghrelin gene expression in the mouse stomach.
Control water, or H2 water (0.8 ml/mouse) made with a stick of magnesium (about 0.04 mM H2; see Methods), was administered to mice (41–48 weeks of age; n = 4 per group) once a day for 4 days. Three hr after the final ingestion, the stomach was removed and prepared for qPCR. Data were normalized with respect to expression of RPL4 mRNA and are represented as mean ± SEM.
Figure 2
Figure 2. Plasma ghrelin levels following administration of oral H2 water.
Mice (16–21 weeks, average 18.5 weeks, n = 5–10) received H2 water (or control water) as in Fig. 1 for 1, 2 or 4 d. Blood was collected on the final day of the experiment, 4 hr after the last ingestion, and plasma levels of ghrelin were quantified by ELISA. Panel shows data from male mice. The ghrelin level was significantly higher only in the group of mice drinking H2 water for 4 d. Data are represented as mean ± SEM.
Figure 3
Figure 3. H2 water increases plasma ghrelin levels in a β1-adrenergic receptor-dependent manner.
Plasma ghrelin levels were measured on the last day of experiment. Each group of mice (11–13 weeks of age; n = 4–5 for each group) was administered control water or H2 water (made with an open-air water electrolysis system) once a day for 4 d with or without i.p. injection of atenolol (10 mg/kg) prior to water ingestion. The H2 water group showed a significant increase in plasma ghrelin level compared to the control group (* p = 0.038), which was abrogated by pretreatment with atenolol (** p = 0.039). Datafare represented as mean ± SEM.
Figure 4
Figure 4. Inhibition of ghrelin secretion or the ghrelin receptor antagonist cancels the neuroprotective effect of oral H2 water in MPTP-induced Parkinson's disease model mice (male, 8–12 weeks, n = 2–5 for each group).
Saturated H2 water was made daily as described in materials and methods. (a): Tyrosine hydroxylase (TH) staining was performed in substantia nigra pars compacta from saline-injected (i–iv), D-Lys3 GHRP-6-injected (v–viii), and atenolol-injected (ix–xii) mice, supplied with either normal tap water (control) or H2 water for 7 days. MPTP was injected (i.p.) after 7 days of D-Lys3 GHRP-6, atenolol, or saline-injection (ii, iv, vi, viii, x, and xii). (b): Summary of the immunoblotting analysis of TH protein in the substantia nigra tissue (n = 3–6 for each group) and the representative blot images were shown underneath (two samples per group). The TH band intensity was normalized to the band of actin on the same sample. (## p = 0.007, i vs. ii; 0.010, ii vs. iv; 0.004, vi vs. vii; # p = 0.043, iv vs. viii; 0.023, v vs. vi). Data in figures are represented as mean ± SEM. (c): Summary of the stereological analysis of nigral dopaminergic neurons. Although MPTP injections caused significant loss of TH-positive neurons (ii) (** p = 0.0001, i vs. ii), drinking H2 water for 7 days prior to MPTP-treatment significantly attenuates the loss of TH-positive cells (iv) (# p = 0.008, ii vs. iv). D-Lys3 GHRP-6, growth hormone secretagogue receptor antagonist, or β1-adrenoceptor blocker, atenolol, canceled the preservation effect of oral H2 water (viii and xii) (# p = 0.008; ii vs. iv; p = 0.003, iv vs. viii; p = 0.0003, iv vs. xii). Data in figures are represented as mean ± SEM. The statistical significance of data was assessed by one-way ANOVA followed by Benferroni test.

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