Mapping dopaminergic deficiencies in the substantia nigra/ventral tegmental area in schizophrenia

Abstract

Previous work from our laboratory showed deficits in tyrosine hydroxylase protein expression within the substantia nigra/ventral tegmental area (SN/VTA) in schizophrenia. However, little is known about the nature and specific location of these deficits within the SN/VTA. The present study had two aims: (1) test if tyrosine hydroxylase deficits could be explained as the result of neuronal loss; (2) assess if deficits in tyrosine hydroxylase are sub-region specific within the SN/VTA, and thus, could affect specific dopaminergic pathways. To achieve these objectives: (1) we obtained estimates of the number of dopaminergic neurons, total number of neurons, and their ratio in matched SN/VTA schizophrenia and control samples; (2) we performed a qualitative assessment in SN/VTA schizophrenia and control matched samples that were processed simultaneously for tyrosine hydroxylase immunohistochemistry. We did not find any significant differences in the total number of neurons, dopaminergic neurons, or their ratio. Our qualitative study of TH expression showed a conspicuous decrease in labeling of neuronal processes and cell bodies within the SN/VTA, which was sub-region specific. Dorsal diencephalic dopaminergic populations of the SN/VTA presented the most conspicuous decrease in TH labeling. These data support the existence of pathway-specific dopaminergic deficits that would affect the dopamine input to the cortex without significant neuronal loss. Interestingly, these findings support earlier reports of decreases in tyrosine hydroxylase labeling in the target areas for this dopaminergic input in the prefrontal and entorhinal cortex. Finally, our findings support that tyrosine hydroxylase deficits could contribute to the hypodopaminergic state observed in cortical areas in schizophrenia.

Keywords: Immunohistochemistry; Mesocortical; Mesolimbic; Neuropathology; Neuropsychiatric disorders; Tyrosine hydroxylase.

Figure 1. Neuronal counts for the entire SN/VTA

A–B) Number of total and dopaminergic neurons in the schizophrenia and control groups. In (A), data are presented as the mean neuronal count for each group with their standard deviation. In (B), individual data for each subject is plotted, with the horizontal bar indicating the mean for the group. C–D) Ratio of dopaminergic/total neurons. The bar graph in (C) represents the mean ratio for each group and their standard deviations. The scatter plot in (D) represents the ratio values for each individual subject, with the horizontal bar indicating the mean ratio for each group. No significant differences were observed for any of these measures between the two groups for the SN/VTA as a whole. Abbreviations: CTRL: control; SZ: schizophrenia

Figure 2. Neuronal counts in the diencephalic (rostral) SN/VTA

A–B) Number of total and dopaminergic neurons in the schizophrenia and control groups for the diencephalic sub-region of the SN/VTA. In (A), data are presented as the mean neuronal count for each group with their standard deviation. In (B), individual data for each subject is plotted, with the horizontal bar indicating the mean for the group. C–D) Ratio of dopaminergic/total neurons. The bar graph in (C) represents the mean ratio for each group and their standard deviations. The scatter plot in (D) represents the ratio values for each individual subject, with the horizontal bar indicating the mean ratio for each group. No significant differences were observed for any of these measures between the two groups in the diencephalic SN/VTA. Abbreviations: CTRL: control; SZ: schizophrenia

Figure 3. Neuronal counts in the mesencephalic (mid-caudal) SN/VTA

A–B) Number of total and dopaminergic neurons in the schizophrenia and control groups for the mesencephalic sub-region of the SN/VTA. In (A), data are presented as the mean neuronal count for each group with their standard deviation. In (B), individual data for each subject is plotted, with the horizontal bar indicating the mean for the group. C–D) Ratio of dopaminergic/total neurons. The bar graph in (C) represents the mean ratio for each group and their standard deviations. The scatter plot in (D) represents the ratio values for each individual subject, with the horizontal bar indicating the mean ratio for each group. No significant differences were observed for any of these measures between the two groups in the mesencephalic SN/VTA. Abbreviations: CTRL: control; SZ: schizophrenia

Figure 4. Tyrosine hydroxylase labeling in the diencephalic SN/VTA

A) Sagittal schematic drawing indicating the rostro-caudal level of sectioning (red line) of the images shown in this figure. B) Representative schematic drawing of a transverse section at the level shown in A. Three SN/VTA subareas defined by the dorso-ventral and medio-lateral location of the dopaminergic neurons are indicated (dashed lines in gray-shaded area). There was a conspicuous qualitative difference in overall immunolabeling between matched schizophrenia (C–E) and control (F–H) samples in the diencephalic sub-region of the SN/VTA. Note also that within this sub-region, the dorsal area of the SN/VTA presented the weakest immunolabeling in the schizophrenia samples (C), while was very intensely labeled in the controls (F). Ventro-lateral areas of the SN/VTA presented also notably weaker immunolabeling in schizophrenia samples (D) compared to the controls (G). Differences in labeling were less apparent when comparing the ventro-medial area of matched schizophrenia (E) and control samples (H). The white asterisks indicate a similar level within the section for comparison of TH labeling between the schizophrenia and control samples. Coordinates indicate the orientation of the sections: C: caudal; D: dorsal; L: lateral; R: rostral. Dashed lines in D, G indicate the approximate transition between the dorsal (d) and ventro-lateral (vl) regions of the SN/VTA Abbreviations: cp: cerebral peduncle; CTRL: control; d: dorsal SN/VTA; III: oculomotor nerve exit; m: mammillary body; pg: periaqueductal grey; RN: red nucleus; SN/VTA: Substantia nigra/ventral tegmental area; SZ: schizophrenia; Thal: thalamus; vl: ventro-lateral SN/VTA; vm: ventro-medial SN/VTA. Calibration bar: 1 mm in C–H.

Figure 5. Tyrosine hydroxylase labeling of cell and processes in the diencephalic SN/VTA

A, D) Dorsal subarea of the diencephalic SN/VTA: In schizophrenia (A, A′) dopaminergic cell bodies presented the typical neuromelanin pigmentation (n), but TH labeling was very weak. Dopaminergic processes also presented weak labeling for TH (white arrowheads). The matched control (D, D′) displayed very intense immunostaining for TH both in cell bodies and processes. B, E) The ventro-lateral subarea of the diencephalic SN/VTA also presented conspicuous deficits in TH immunolabeling in schizophrenia (B, B′) compared with a matched control (E, E′). Note also that there is a gradient of TH immunolabeling deficit, with the areas close to the transition between the ventro-lateral (vl) and dorsal (d) subareas presenting even weaker TH labeling in the schizophrenia (B) but not in the control (E). Boxed areas in A, D, B, E are shown at higher magnification in A′, D′, B′, E′, respectively. C, F) The ventro-medial subarea of the SN/VTA presented scarce dopaminergic neurons (arrows) sparsely distributed among a dense net of thin TH labeled processes both in the schizophrenia (C) and control (F) samples. The differences in TH immunolabeling between schizophrenia and matched controls for this ventro-medial subarea were more subtle, displaying overall moderate qualitative deficits of TH labeling in their dense net of thin dopaminergic processes, and no clear qualitative reductions of labeling in dopaminergic cell bodies. The white asterisks indicate a similar level within the section for comparison of TH labeling between the schizophrenia and control samples. Coordinates indicate the orientation of the sections: D: dorsal; L: lateral. Abbreviations: CTRL: control; SZ: schizophrenia. Calibration bar: 250 μm in A–F.

Figure 6. Tyrosine hydroxylase labeling in the mesencephalic SN/VTA

A) Sagittal schematic drawing indicating the rostro-caudal level of sectioning (red line) of the images shown in this figure. B) Representative schematic drawing of a transverse section at the level shown in A. Three SN/VTA subareas defined by the dorso-ventral and medio-lateral location of the dopaminergic neurons are indicated (dashed lines in gray-shaded area). C, F) These images show the overall pattern of TH labeling in the mesencephalic sub-region of matched schizophrenia (C) and control (F) samples. In these low magnification images the differences in labeling between schizophrenia and controls were very subtle. D, G) At higher magnification, in the dorsal (d) subarea there were moderate-to-subtle deficits in labeling in schizophrenia (D) compared with matched control (G) samples. E, H) In the ventro-lateral (vl) subarea there were no obvious differences between schizophrenia (E) and matched control (H) samples. Coordinates indicate the orientation of the sections: C: caudal; D: dorsal; L: lateral; R: rostral. The white asterisks in C and F indicate a similar level within the section for comparison of TH labeling between the schizophrenia and control samples Dashed lines in C, F indicate the approximate transition between the dorsal (d) and ventro-lateral (vl) regions of the SN/VTA. The areas marked with dashed boxes in C and F are shown at high magnification in D, E, G, H. Arrowheads in D–E and G–H indicate some examples of TH immunolabeling in thin processes. Abbreviations: cp: cerebral peduncle; CTRL: control; d: dorsal SN/VTA; III: oculomotor nerve exit; m: mammillary body; pg: periaqueductal grey; RN: red nucleus; SN/VTA: Substantia nigra/ventral tegmental area; SZ: schizophrenia; Thal: thalamus; vl: ventro-lateral SN/VTA; vm: ventro-medial SN/VTA. Calibration bars: 1 mm in C, F; 500 μm in D, E, G, H.