Cholinergic and GABAergic Inputs Drive Patterned Spontaneous Motoneuron Activity before Target Contact

Abstract

Patterned spontaneous electrical activity has been demonstrated in a number of developing neural circuits and has been proposed to play a role in refining connectivity once axons reach their targets. Using an isolated spinal cord preparation, we have found that chick lumbosacral motor axons exhibit highly regular bursts of activity from embryonic day 4 (E4) (stage 24-25), shortly after they exit the spinal cord and while still en route toward their target muscles. Similar bursts could be evoked by stimulating descending pathways at cervical or thoracic levels. Unlike older embryonic cord circuits, the major excitatory transmitter driving activity was not glutamate but acetylcholine, acting primarily though nicotinic non-alpha7 receptors. The circuit driving bursting was surprisingly robust and plastic, because bursting was only transiently blocked by cholinergic antagonists, and following recovery, was now driven by GABAergic inputs. Permanent blockade of spontaneous activity was only achieved by a combination of cholinergic antagonists and bicuculline, a GABAA antagonist. The early occurrence of patterned motor activity suggests that it could be playing a role in either peripheral pathfinding or spinal cord circuit formation and maturation. Finally, the characteristic differences in burst parameters already evident between different motoneuron pools at E4 would require that the combination of transcription factors responsible for specifying pool identity to have acted even earlier.

Fast noninvasive activation and inhibition of neural and network activity by vertebrate rhodopsin and green algae channelrhodopsin

“…4 A and B) revealed that as in control embryos the electroporated embryos exhibited episodes consisting of several bursts every 4 min (Fig. 4 B and C) (25). Thus the electroporation protocol and expression of these constructs over several days did not appear to have any adverse effects on the development of the cord circuits responsible to generating this activity.…”

“…4 E and F). Thus light, as has been previously shown for electrical stimulation (24,25), can be used to elicit episodes either by activation of local lumbar interneurons and motoneurons or activation of neurons many cord segments distant.…”

“…Our next goal was to show that these light-sensitive proteins could be used to control circuit behavior in whole animal preparations. Early embryonic chick spinal cords exhibit rhythmic episodes of spontaneous bursting activity, which are generated by recurrent excitatory connections between motoneurons and GABAergic and glycinergic interneurons, all of which are excitatory at this stage of development (23)(24)(25). Recently, it has been shown that the normal pattern and frequency of this early spontaneous activity is required for appropriate motor axon pathfinding in the chick (14) and for the development of cord circuits that enable appropriate flexorextensor and right-left phasing during locomotor-like activity in the mouse (26).…”

Acetylcholine negatively regulates development of the neuromuscular junction through distinct cellular mechanisms

“…For example, the neurotransmitter acetylcholine (ACh) is released from embryonic motor neurons (MNs) at the neuromuscular junction (NMJ), and it negatively regulates survival, axon branching, and synapse formation (4)(5)(6)(7). ACh is also released from developing neurons even before they arrive at their target, and it acts in an autocrine and/or paracrine fashion to regulate growth (8), pathfinding (9), spontaneous activity (10), and target selection (11). Whereas the NMJ has served as an excellent model to elucidate the process of synapse formation, the cellular and molecular mechanisms by which ACh regulates this process are still unclear; this is in part because the expression of ACh receptor (AChR) subtypes varies over space and time during the development of this synapse.…”

Onset of ETS expression is not accelerated by premature exposure to signals from limb mesenchyme

“…Although all these motor neurons originate from a common precursor population in ventral neural tube and share many features in common at maturity, each motor pool has unique distinguishing characteristics. Neurons within an individual motor pool reside in a characteristic position in the LMC (Landmesser, 1978), exhibit a distinctive pattern of spontaneous electrical activity (Milner and Landmesser, 1999), express a unique set of LIM homeodomain and ETS transcription factors and cadherins (Tsuchida et al, 1994;Lin et al, 1998;Price et al, 2002), innervate a single target muscle, and receive monosynaptic input from sensory neurons supplying the same muscle (reviewed in Chen et al, 2003). These distinguishing properties of motor pools arise through the sequential activation of different transcriptional networks (reviewed in Shirasaki and Pfaff, 2002).…”