Dopamine D1, D2, D3 receptors, vesicular monoamine transporter type-2 (VMAT2) and dopamine transporter (DAT) densities in aged human brain

Abstract

The dopamine D(1), D(2), D(3) receptors, vesicular monoamine transporter type-2 (VMAT2), and dopamine transporter (DAT) densities were measured in 11 aged human brains (aged 77-107.8, mean: 91 years) by quantitative autoradiography. The density of D(1) receptors, VMAT2, and DAT was measured using [(3)H]SCH23390, [(3)H]dihydrotetrabenazine, and [(3)H]WIN35428, respectively. The density of D(2) and D(3) receptors was calculated using the D(3)-preferring radioligand, [(3)H]WC-10 and the D(2)-preferring radioligand [(3)H]raclopride using a mathematical model developed previously by our group. Dopamine D(1), D(2), and D(3) receptors are extensively distributed throughout striatum; the highest density of D(3) receptors occurred in the nucleus accumbens (NAc). The density of the DAT is 10-20-fold lower than that of VMAT2 in striatal regions. Dopamine D(3) receptor density exceeded D(2) receptor densities in extrastriatal regions, and thalamus contained a high level of D(3) receptors with negligible D(2) receptors. The density of dopamine D(1) linearly correlated with D(3) receptor density in the thalamus. The density of the DAT was negligible in the extrastriatal regions whereas the VMAT2 was expressed in moderate density. D(3) receptor and VMAT2 densities were in similar level between the aged human and aged rhesus brain samples, whereas aged human brain samples had lower range of densities of D(1) and D(2) receptors and DAT compared with the aged rhesus monkey brain. The differential density of D(3) and D(2) receptors in human brain will be useful in the interpretation of PET imaging studies in human subjects with existing radiotracers, and assist in the validation of newer PET radiotracers having a higher selectivity for dopamine D(2) or D(3) receptors.

Figure 1. Chemical structures of [³H]WC-10 and [³H]raclopride.

K_d values were obtained through saturation binding of [³H]WC-10 and [³H]raclopride to cloned human D₃ and D_2L receptors expressed in HEK cells. a₁, and b₁ represent the fractional receptor occupancy to dopamine D₂ and D₃ receptors in human brain at a ligand concentration of 3.54 nM for [³H]WC-10. a₂ and b₂ represent the same parameters at a ligand concentration of 2.50 nM [³H]raclopride. The receptor occupancy fractions were calculated from the saturation binding isotherm using the K_d values. *Data were taken from Xu et al. (2009).

Figure 2. Quantitative autoradiographic analysis of dopamine receptors, DAT and DTBZ densities in the precommissural striatal regions.

Autoradiograms show total binding of 1.44 nM [³H]SCH23390, 2.50 nM[³H]raclopride, 3.54 nM [³H]WC-10, 2.19 nM [³H]WIN35428, and 4.53 nM [³H]DTBZ (A), and nonspecific binding in presence of 1 µM (+) butaclamol (for [³H]SCH23390), 1 µM S(-)-eticlopride (for [³H]raclopride and [³H]WC-10), 1 µM nomifensine (for WIN35428) and 1 µM S(-)-tetrabenazine (for DTBZ) (B) in the precommissural striatal regions of human brain sections. The adjacent section shows cresyl violet staining to identify related anatomical structures (C). [³H]Microscale standards (ranging from 0 to 36.3 nCi/mg) were also counted (D). Quantitative analysis of dopamine D₁, D₂, and D₃ receptors, and DAT and DTBZ densities (fmol/mg) and the dopamine D₂ ∶ D₃ receptor density ratio in human striatal regions are shown in E and F respectively. The numbers 1 through 4 designate the following CNS anatomical regions: 1: Precommissural Putamen (PrePu); 2: Precommissural caudate (PreCd);3: Nucleus accumbens (NAc); 4: Internal capsule (IC). *p<0.05, #p<0.01 compared to NAc.

Figure 3. Quantitative autoradiographic analysis of dopamine D₃ receptors, DAT and DTBZ densities in the globus pallidus.

Autoradiograms show total binding of 1.44 nM [³H]SCH23390, 2.50 nM [³H]raclopride, 3.54 nM [³H]WC-10, 2.19 nM [³H]WIN35428, 4.53 nM [³H]DTBZ (A), and nonspecific binding in presence of 1 µM (+) butaclamol (for [³H]SCH23390), 1 µM S(-)-eticlopride (for [³H]raclopride and [³H]WC-10), 1 µM nomifensine (for [³H]WIN35428) and 1 µM S(-)-tetrabenazine (for [³H]DTBZ) (B) in the globus pallidus of aged human brain sections. The adjacent section shows cresyl violet staining to identify related anatomical structures (C). [³H]Microscale standards (ranging from 0 to 36.3 nCi/mg) were also counted (D). Quantitative analysis of dopamine D₁, D₂, and D₃ receptors, DAT and DTBZ densities (fmol/mg) and the dopamine D₂ ∶ D₃ receptor density ratio in human globus pallidus are shown in E and F, respectively. The numbers 1 through 5 designate the following CNS anatomical regions: 1: Putamen; 2: Caudate; 3: Globus pallidus external part (GPe); 4: Globus pallidus internal part (GPi); 5: Internal capsule (IC). *p<0.05 compared to GPe.

Figure 4. Quantitative autoradiographic analysis of dopamine receptors, DAT and DTBZ densities in the thalamus.

Autoradiograms show total binding of 1.44 nM [³H]SCH23390, 2.50 nM [³H]raclopride, 3.54 nM [³H]WC-10, 2.19 nM [³H]WIN35428, 4.53 nM [³H]DTBZ (A), and nonspecific binding in presence of 1 µM (+) butaclamol (for [³H]SCH23390), 1 µM S(-)-eticlopride (for [³H]raclopride and [³H]WC-10), 1 µM nomifensine (for WIN35428) and 1 µM S(-)-tetrabenazine (for DTBZ) (B) in the thalamus of human brain sections. The adjacent section shows cresyl violet staining to identify related anatomical structures (C). [³H]Microscale standards (ranging from 0 to 36.3 nCi/mg) were also counted (D). Quantitative analysis of dopamine D₁, D₂, and D₃ receptors, DAT and DTBZ densities (fmol/mg) and the dopamine D₂ ∶ D₃ receptor density ratio in human brain are shown in E and F, respectively. Linear correlation analysis of the average dopamine D₁ and D₃ receptor densities in human thalamus is shown in (G). The numbers 1 through 4 designate the following CNS anatomical regions: 1: Postcommissural putamen (PostPu); 2: Postcommissural caudate (PosCd); 3: Thalamus; 4: Internal capsule (IC). #p<0.01 compared to thalamus.

Figure 5. Quantitative autoradiographic analysis of dopamine receptors, and DAT and DTBZ densities in the substantia nigra.

Autoradiograms show total binding of 1.44 nM [³H]SCH23390, 2.50 nM[³H]raclopride, 3.54 nM [³H]WC-10, 2.19 nM [³H]WIN35428, 4.53 nM [³H]DTBZ (A), and nonspecific binding in presence of 1 uM (+) butaclamol (for [³H]SCH23390), 1 µM S(-)-eticlopride (for [³H]raclopride and [³H]WC-10), 1 µM nomifensine (for WIN35428) and 1 µM S(-)-tetrabenazine (for DTBZ) (B) in the substantia nigra (SN) of aged human brain sections. [³H]Microscale standards (ranging from 0 to 36.3 nCi/mg) were also counted (C). Quantitative analysis of dopamine D₁, D₂ and D₃ receptors, and DAT and DTBZ densities (fmol/mg) and the dopamine D₂ ∶ D₃ receptor density ratio in human SN and red nucleus are shown in D and E respectively. The numbers 1 through 3 designate the following CNS anatomical regions: 1: Substantia nigra (SN); 2: Red nucleus (RN); 3: Thalamus.

Figure 6. Comparison of dopamine D₁, D₂, and D₃ receptors, and DAT and DTBZ densities in the striatal regions between an aged rhesus monkey (25 years old) and aged human brain samples.

Autoradiograms show neuroanatomical localization of [³H]SCH23390 for D₁, [³H]raclopride for D₂, [³H]WC-10 for D₃ receptors, [³H]WIN35428 for DAT and [³H]DTBZ for VMAT2 in the striatal regions of rhesus monkey (A) and aged human brain (B). [³H]Microscale stnadards (ranging from 0 to 36.3 nCi/mg) (C). The numbers 1 through 8 in panels (A) (B) designate the following CNS anatomical regions: 1: Monkey putamen; 2: Monkey caudate; 3: Monkey globus pallidus; 4: Monkey thalamus; 5: Human putamen; 6: Human caudate; 7: Human globus pallidus; 8: Human thalamus.

Figure 7. Correlation of DAT with VMAT2 in the striatal regions and substantia nigra.

The correlation between the VMAT2 and DAT densities in the precommissural putamen (PrePu), caudate (PreCd) and substantia nigra (SN) (A). Correlation of the VMAT densities between the substantia nigra (SN) and PrePu or PreCd (B). Correlation of the DAT densities between the SN and PrePu or PreCd (C). The average VMAT DAT density ratio in the PrePu, PreCd and SN (D). #p<0.01 compared to SN.