Dopaminergic projections from midbrain to primary motor cortex mediate motor skill learning

Abstract

The primary motor cortex (M1) of the rat contains dopaminergic terminals. The origin of this dopaminergic projection and its functional role for movement are obscure. Other areas of cortex receive dopaminergic projections from the ventral tegmental area (VTA) of the midbrain, and these projections are involved in learning phenomena. We therefore hypothesized that M1 receives a dopaminergic projection from VTA and that this projection mediates the learning of a motor skill by inducing cellular plasticity events in M1. Retrograde tracing from M1 of Long-Evans rats in conjunction with tyrosine hydroxylase immunohistochemistry identified dopaminergic cell bodies in VTA. Electrical stimulation of VTA induced expression of the immediate-early gene c-fos in M1, which was blocked by intracortical injections of D(1) and D(2) antagonists. Destroying VTA dopaminergic neurons prevented the improvements in forelimb reaching seen in controls during daily training. Learning recovered on administration of levodopa into the M1 of VTA-lesioned animals. Lesioning VTA did not affect performance of an already learned skill, hence, left movement execution intact. These findings provide evidence that dopaminergic terminals in M1 originate in VTA, contribute to M1 plasticity, and are necessary for successful motor skill learning. Because VTA dopaminergic neurons are known to signal rewards, the VTA-to-M1 projection is a candidate for relaying reward information that could directly support the encoding of a motor skill within M1.

Figure 1.

Retrograde tracing from M1 identifies dopaminergic neurons in the VTA and SN. a, Representative images from VTA (magnification, 20×). The white arrowheads indicate dopaminergic neurons (left, anti-TH), neurons projecting to M1 (middle, retrograde Fast Blue labeling), double-labeled neurons (right, anti-TH and Fast Blue). The open arrowheads indicate a non-dopaminergic neuron (anti-TH negative) projecting to M1 (Fast Blue-positive). Scale bar, 30 μm. b, Superposition of double-labeled neurons (magenta) and Fast Blue-labeled neurons (blue) in three representative sections (positions relative to bregma) derived from six animals. ▾, Tracer-injected hemisphere; APN, anterior pretectal nucleus; DMN, deep mesencephalic nucleus; MCPC, magnocellular nucleus of posterior commissure; mp, mamillary peduncle; PR, prerubral field. Scale bars, 1 mm. c, Average number of double-labeled neurons in VTA and SN for selected sections (position relative to bregma; error bars indicate SEM).

Figure 2.

VTA stimulation induces c-fos expression in M1 via release of dopamine. a, Representative image of M1 demonstrating c-fos expression in response to ipsilateral VTA stimulation (VTA Stim) and unstimulated control (Sham). Images were taken from the right hemisphere, 2.5 mm anterior to bregma. Scale bars, 250 μm. b, Representative images of c-fos expression patterns in M1 after VTA stimulation combined with either intracortical vehicle (VTA Stim + vehicle) or D₁ plus D₂ antagonist injections (VTA Stim + D1/2 ant). Images were taken from the right hemisphere at 2.5 mm anterior to bregma. Scale bars, 250 μm.

Figure 3.

Destroying dopaminergic neurons in VTA depletes dopaminergic terminals in M1. a, Representative TH immunofluorescence staining of M1 shows a clear reduction of TH-positive dopaminergic terminals throughout all cortical layers in 6-OHDA-lesioned animals. Scale bar, 250 μm. b, Example of TH immunohistochemistry shows effective destruction of dopaminergic neurons in the VTA after 6-OHDA injection in conjunction with desipramine intraperitoneally to protect noradrenergic neurons (right) compared with the vehicle-injected VTA (left). The arrow indicates injection site. Scale bar, 50 μm. c, Western blot analysis of M1 TH immunoreactivity demonstrates quantitatively the loss of dopaminergic neurons after 6-OHDA lesions in VTA compared with vehicle-injected controls (n = 8; *p = 0.048). GAPDH was used as a positive control protein and remained unchanged after 6-OHDA lesions. Error bars indicate SEM.

Figure 4.

Intact dopaminergic projection from VTA to M1 is required for motor skill learning. a, Lesioning VTA prevented training-induced improvements in reaching performance (lesion_1 and lesion_2) compared with sham-lesioned controls (days 1–8; white background). Infusing levodopa directly into M1 (days 9–16; gray background) rescued the VTA lesion-induced deficit (lesion_1; n = 7), whereas there was no such restoration on vehicle injection (lesion_2; n = 9) and no additional improvement by levodopa injections in rats that had already acquired the task (sham; n = 8). Lesion_1 animals maintained performance after discontinuation of levodopa substitution (days 17–24; gray-striped background). b, Short-term improvement in session 1 is maintained after lesioning VTA (blue) compared with sham (black). c, Destroying dopaminergic neurons in VTA after 8 d of training (plateau phase without additional learning; n = 6) does neither affect performance indicating unaffected movement execution, nor latency. d, Latencies between pellet removal and subsequent door opening, an index of motivation, were not affected by VTA lesions. e, Rotarod performance, an index of general motor function, decreased because of surgical instrumentation but not because of 6-OHDA injections into VTA. Error bars indicate SEM.