Browsing by Subject "Midbrain"

The development and function of the nervous system relies on complex regulation of gene expression programs. MicroRNAs (miRNAs) are small RNAs that have diverse functions in mammalian development and disease. In concert with the RNA-induced silencing complex, miRNAs repress translation by binding to target mRNAs. The nervous system contains the largest proportion of miRNAs, yet few have been functionally characterized in vivo.

miR-133b is a highly conserved miRNA embedded in the sequence of 7H4, a noncoding RNA that is enriched at the neuromuscular junction (NMJ), a large synapse that is essential for eliciting muscle contraction and movement. I have found that, like 7H4, miR-133b expression is enriched at the NMJ and upregulated postnatally, coinciding with important events in synaptic maturation, including synaptic growth and elimination. Knockdown of miR-133b in postnatal muscle by electroporation of modified antisense oligonucleotides gave rise to abnormally large synapses, indicating a role for miR-133b in synaptic maturation. To specifically remove miR-133b in vivo, I generated a mouse containing a targeted deletion of the miR-133b stemloop. NMJ maturation and synapse elimination proceeded normally in miR-133b knockout mice, suggesting that miR-133b may have other functions at the synapse. The expression of 7H4 and miR-133b is upregulated following nerve transection, consistent with a role in synaptic regeneration. Indeed, NMJ reinnervation is delayed in miR-133b KO mice following nerve crush, but not nerve cut. These data suggest that miR-133b may have a specific protective function at the synapse that could be relevant to disease states, including amyotrophic lateral sclerosis (ALS), where NMJ denervation occurs following motor neuron cell death. However, loss of miR-133b did not affect survival or disease progression in the SOD1(G93A) mouse model, differentiating its role from that of miR-206, another miRNA found in 7H4.

miR-133b has recently been proposed to regulate the development and maintenance of midbrain dopaminergic (mDA) neurons. mDA neurons have critical functions in the control of movement and emotion, and their degeneration leads to motor and cognitive defects in Parkinson's disease. miR-133b is enriched in the midbrain and regulates mDA neuron differentiation in vitro by targeting Pitx3, a transcription factor required for appropriate development of substantia nigra DA neurons. However, the function of miR-133b in the intact midbrain has not been determined. miR-133b KO mice have normal numbers of midbrain dopaminergic neurons during development and aging. Moreover, dopamine neurotransmitter levels are unchanged in the striatum and other brain regions, while expression of dopaminergic genes including Pitx3 is also unaffected. Finally, miR-133b null mice display normal motor coordination and activity, suggesting that miR-133b does not play a significant role in the development or maintenance of the mDA neuron population.

Many of the skills we value most as humans, such as speech and learning to play musical instruments, are learned in the absence of external reinforcement. However, the model systems most commonly used to study motor learning employ learning paradigms in which animals perform behaviors in response to external rewards or punishments. Here I use the zebra finch, an Australian songbird that can learn its song as a juvenile in the absence of external reinforcement as well as modify its song in response to external cues as an adult, to study the circuit mechanisms underlying both internally and externally reinforced forms of learning. Using a combination of intersectional genetic and microdialysis techniques, I show that a striatonigral pathway and its downstream effectors, namely D1-type dopamine receptors, are necessary for both internally reinforced juvenile learning and externally reinforced adult learning, as wells as for song modification in response to social cues or to deafening. In addition, I employ optogenetic stimulation during singing to demonstrate that this striatonigral projection is sufficient to drive learning. Interestingly, I find that neither the striatonigral pathway nor D1-type dopamine receptors are necessary for recovery of pitch after externally driven pitch learning. In all, I establish that a common mechanism underlies both internally and externally reinforced vocal learning.

Introduction: Intervertebral disc herniation may contribute to nerve root compression or inflammatory processes that are associated with radicular pain and motor deficits. Molecular changes at the affected dorsal root ganglion (DRG), spinal cord, and even midbrain, have been documented in rat models of radiculopathy or nerve injury. The objective of this study was to evaluate gait mechanics and the expression of key pain receptors in the midbrain of rats after induced radiculopathy in order to test the hypothesis that DRG injury can promote molecular changes in the midbrain. Materials and Methods: Radiculopathy was induced by harvesting tail nucleus pulposus (NP) and placing upon the right L5 DRG in Sprague-Dawley rats. Tail nucleus pulposus (NP) was harvested and discarded in sham-operated rats. At 1 and 4 weeks after surgery, DRGs were sectioned and tested for immunoreactivity to astrocytes and microglial. Also at 1 and 4 weeks after surgery, midbrains were sectioned and tested for immunoreactivity to serotonin (5HT2B), mu-opioid (μ-OR), and metabotropic glutamate (mGluR4 and 5) receptor antibodies. Quantitative analysis was performed on all midbrain immunostained images and compared to naïve controls. Cerebral spinal fluid was also extracted at 1 and 4 weeks after surgery for monocyte-chemoattractant protein (MCP-1) assessment. Results: NP-treated animals placed less weight on the affected limb 1 week after surgery and experienced mechanical hypersensitivity over the entire time of the study. Astroctye activation was observed at the DRG 4 weeks after surgery. An increased expression of 5HT2B was observed in NP-treated rats at 1, but not at 4 weeks. Increased expression of μ-OR and mGluR5 was observed in the periaqueductal gray (PAG) region of NP-treated rat midbrains at 1 and 4 weeks post-surgery. By contrast, increased expression levels of mGluR5 in the PAG region of sham animals reverted to naïve levels by 4 weeks after surgery. No changes were observed in expression levels of mGluR4 in either sham or NP-treated animals at any point in this study. MCP-1 levels were higher in NP-treated animals at 4 weeks compared to sham animals. Conclusion: These observations support the hypothesis that the midbrain responds to injury at the DRG with a transient and adaptive change in receptors regulating pain mechanisms.