Evidence of COMT dysfunction in the olfactory bulb in Parkinson's disease

Abstract

Hyposmia is one of the most prevalent non-motor symptoms of Parkinson's disease and antecedes motor dysfunction by up to a decade. However, the underlying pathophysiology remains poorly understood. In this study, we investigated the mechanisms of dopamine metabolism in post-mortem olfactory bulbs from ten Parkinson's disease and ten neurologic control subjects. In contrast to the loss of dopaminergic neurons in the midbrain, we observed an increase in tyrosine hydroxylase-positive neurons in the Parkinson's disease olfactory bulb, suggesting a potential role for dopamine in the hyposmia associated with the condition. Using immunohistochemistry, high-performance liquid chromatography, western blot, and enzyme-linked immunosorbent assays, we demonstrate a reduction in catechol-O-methyltransferase catabolism of dopamine to homovanillic acid, potentially due to a depletion of the methyl donor substrate S-adenosyl methionine. We hypothesized that reduction in catechol-O-methyltransferase activity would result in increased dopamine occupation of the D₂ receptor, and consequent inhibition of olfactory processing. Next, we conducted pharmacological interventions to modify dopamine dynamics in hyposmic tau knockout mice, which exhibit altered dopamine metabolism. Our hypothesis was supported by the observation that the D₂ receptor antagonist haloperidol temporarily alleviated olfactory deficits in these tau knockout mice. This study implicates a potential role of catechol-O-methyltransferase-mediated dopamine metabolism in the early olfactory impairments associated with Parkinson's disease.

Keywords: Catechol-O-methyltransferase; Dopamine; Hyposmia; Olfaction; Parkinson’s disease.

Fig. 1

Dopamine metabolism is altered in the PD olfactory bulb. a Representative immunoblots of olfactory bulb lysates stained for tyrosine hydroxylase (TH), pTH (ser40) and β-actin. b Quantification of TH and pTH immunoblot densitometry presented as % of neurological control, and ratio of pTH:TH. c Quantification of TH⁺ neurons, representative images of TH-stained olfactory bulb sections from one NC and one PD sample; red arrows indicate TH⁺ neurons, scale bar represents 200 μm. d Dopamine, 3,4-dihydroxyphenylacetic acid (DOPAC), and homovanillic acid (HVA) concentration determined by HPLC-ED. e Ratio of DOPAC:dopamine, HVA:DOPAC and HVA:dopamine in olfactory bulb tissue. f Dopamine turnover (DOPAC + HVA/dopamine). g Proportion of HVA to DOPAC. h Catechol-O-methyltransferase (COMT) protein level, S-adenosylmethionine (SAMe) protein levels, and magnesium levels in tissue lysate. i Monoamine oxidase activity level. j D₂ receptor protein level. NC (N = 10) and PD (N = 10). Analyses performed by Students’ T Test; *P < 0.05, ** P < 0.01, ***P < 0.001, ****P < 0.0001

Fig. 2

Correlation analysis between dopamine synthesis and metabolism in PD olfactory bulb. a Correlation matrix assessing the relation between key dopamine synthesis and metabolism enzymes, dopamine, and dopamine metabolites in neurological control (top) and Parkinson’s disease (bottom). The color scale depicts the strength of the Pearson correlation coefficient (r). A positive correlation (blue) indicates that the variables either increase or decrease in the same direction. A negative correlation (red) indicates that the variables increase or decrease in opposing directions. Asterisks depict the significance of the correlation: * P < 0.05, ** P < 0.01. b Positive correlation of monoamine oxidase (MAO) activity and 3,4-dihydroxyphenylacetic acid (DOPAC). c Correlation of catechol-O-methyltransferase (COMT) and homovanillic acid (HVA) in neurological controls (top) and Parkinson’s disease (bottom). Data passed normality testing (D’Agostino–Pearson test) and correlation was determined by Pearson correlations

Fig. 3

Dopamine metabolism is altered in the tau knockout olfactory bulb. a Quantification of TH and pTH immunoblot densitometry presented as % of neurological control, and ratio of pTH:TH. b Representative immunoblots of olfactory bulb lysates stained for tyrosine hydroxylase (TH), pTH (ser40) and total protein. c Dopamine, 3,4-dihydroxyphenylacetic acid (DOPAC), and homovanillic acid (HVA) concentration determined by HPLC-ED. d Dopamine turnover (DOPAC + HVA/dopamine). e Ratio of DOPAC:dopamine, HVA:DOPAC and HVA:dopamine in olfactory bulb tissue. f Quantification of TH⁺ neurons. g Catechol-O-methyltransferase (COMT) protein level, S-adenosylmethionine (SAMe) protein levels, and magnesium levels in tissue lysate. h Images of TH-stained sections from three WT and three tau KO olfactory bulbs, scale bar represents 500 µm; red arrows on 100X images indicate TH+ neurons. Analyses performed by Students’ T test; *P < 0.05, ** P < 0.01

Fig. 4

Dopamine receptor antagonism transiently restores olfactory deficit in tau knockout mice. a ‘Baseline’ odor detection test of WT (N = 29) and tau knockout (N = 31) mice performed at 8-months-old. b Seven days later, animals underwent a ‘test’ ODT 1 h after injection of saline (WT N = 10, tau KO N = 10), 20 mg/kg cocaine (WT N = 9, tau knockout N = 10), or 0.33 mg/kg haloperidol (WT N = 10, tau KO N = 11). c Following a 24-h drug washout period, animals underwent a ‘recovery’ ODT. d Inter-group comparisons of odor detection performance between test days. Analysis was performed by a three-way ANOVA with Sidak post hoc comparisons (denoted by asterisks), *P < 0.05, ****P < 0.0001. Secondary analysis was performed by a one-sample t-test with a hypothetical mean of 50% (denoted by octothorpes). ^#P < 0.05, ^##P < 0.01, ^###P < 0.001, ^####P < 0.0001