Pitx2 cholinergic interneurons are the source of C bouton synapses on brainstem motor neurons

IR and LZ were funded by the European Union, Seventh Framework Programme (FP7/2007–2013), by the European Union and Greek National Funds through the operational program “Education and Lifelong Learning” of the National Strategic Reference Framework, funding program: ARISTEIA II, and by Fondation Santé.Cholinergic neuromodulation has been described throughout the brain and has been implicated in various functions including attention, food intake and response to stress. Cholinergic modulation is also thought to be important for regulating motor systems, as revealed by studies of large cholinergic synapses on spinal motor neurons, called C boutons, which seem to control motor neuron excitability in a task-dependent manner. C boutons on spinal motor neurons stem from spinal interneurons that express the transcription factor Pitx2. C boutons have also been identified on the motor neurons of specific cranial nuclei. However, the source and roles of cranial C boutons are less clear. Previous studies suggest that they originate from Pitx2+ and Pitx2− neurons, in contrast to spinal cord C boutons that originate solely from Pitx2 neurons. Here, we address this controversy using mouse genetics, and demonstrate that brainstem C boutons are Pitx2+ derived. We also identify new Pitx2 populations and map the cholinergic Pitx2 neurons of the mouse brain. Taken together, our data present important new information about the anatomical organization of cholinergic systems which impact motor systems of the brainstem. These findings will enable further analyses of the specific roles of cholinergic modulation in motor control.Publisher PDFPeer reviewe

Glial-derived adenosine modulates spinal motor networks in mice

The activation of purinergic receptors modulates central pattern generators controlling rhythmic motor behaviors, including respiration in rodents and swimming in frog tadpoles. The present study aimed to determine whether purinergic signaling also modulates the mammalian locomotor central pattern generator. This was investigated by using isolated spinal cord preparations obtained from neonatal mice in which locomotor-related activity can be induced pharmacologically. The application of either ATP or adenosine led to a reduction in the frequency of locomotor activity recorded from ventral roots. ATP had no effect when applied in the presence of both the adenosine receptor antagonist theophylline and the ectonucleotidase inhibitor ARL67156, demonstrating that the effects of ATP application result from the breakdown of ATP to adenosine and subsequent activation of adenosine receptors. The application of theophylline or the A1-specific antagonist cyclopentyl dipropylxanthine, but not the A2A-receptor antagonist SCH58261, caused an increase in locomotor burst frequency, demonstrating that endogenously derived adenosine activates A1 receptors during locomotor network activity. Furthermore, theophylline had no effect in the presence of the ectonucleotidase inhibitor ARL67156 or the glial toxins methionine sulfoximine or ethyl fluoracetate, suggesting that endogenous adenosine is derived from ATP, which is released from glia. Finally, adenosine had no effect on slow rhythmic activity recorded upon blockade of all inhibitory transmission, suggesting that adenosine may act via the modulation of inhibitory transmission. Together, these data highlight endogenous purinergic gliotransmission, involving activation of A1 receptors, as an important intrinsic modulatory system controlling the frequency of activity generated by spinal locomotor circuitry in mammals

Backbone–Base Interactions Critical to Quantum Stabilization of Transfer RNA Anticodon Structure

Transfer RNA (tRNA) anticodons adopt a highly ordered 3′-stack without significant base overlap. Density functional theory at the M06-2X/6-31+G­(d,p) level in combination with natural bond orbital analysis was utilized to calculate the intramolecular interactions within the tRNA anticodon that are responsible for stabilizing the stair-stepped conformation. Ten tRNA X-ray crystal structures were obtained from the PDB databank and were trimmed to include only the anticodon bases. Hydrogenic positions were added and optimized for the structures in the stair-stepped conformation. The sugar–phosphate backbone has been retained for these calculations, revealing the role it plays in RNA structural stability. It was found that electrostatic interactions between the sugar–phosphate backbone and the base provide the most stability, rather than the traditionally studied interbase stacking. Base-stacking interactions, though present, were weak and inconsistent. Aqueous solvation was found to have little effect on the intramolecular interactions