Developmental expression of nicotinic receptors in the chick and human spinal cord

Abstract

Naturally occurring programmed cell death of lumbar motor neurons in the chick spinal cord occurs between embryonic day (E) 6 and E12; whereas, a peak of motor neuron degeneration in the human spinal cord occurs between 12 and 16 weeks gestation. One of the major neurotransmitters, acetylcholine, is released from the embryonic motor neuron early in development and is thought to be responsible for early muscle activity that serves as a signal for regulating motor neuron survival. The effects of acetylcholine are mediated by two functionally distinct classes of receptors; namely, muscarinic and nicotinic with nicotinic receptors being used at the neuromuscular synapse. In this study, we determined the developmental expression profile of nicotinic acetylcholine receptor subunits in the chick and human lumbar motor neurons and skeletal muscle using reverse transcription polymerase chain reaction, immunoblots, and immunocytochemistry. Our results show that, in the chick, nicotinic receptor subunits alpha1, alpha4, alpha7, alpha8, and beta2 appear to be regulated during the process of naturally occurring motor neuron cell death in the spinal cord. A new finding was the expression of alpha8 mRNA and protein from E4.5 through E7 in chick motor neurons. Interestingly, we also found that, at E14, alpha8 protein was localized only in sensory dorsal horn neurons. In the developing human spinal cord, we determined that nicotinic receptor subunits alpha1, alpha2, alpha3, alpha4, alpha7, beta2, and beta3 were expressed before the programmed cell death period, and alpha2, alpha4, alpha7, beta2, beta3, and beta4 were expressed during the programmed cell death period. Our data demonstrate that neuronal and muscle nicotinic receptor mRNAs and proteins are expressed during important embryonic periods. This finding raises the possibility that nicotinic receptors play an important role in the spinal cord and skeletal muscle during early development.

The rescue of developing avian motoneurons from programmed cell death by a selective inhibitor of the fetal muscle‐specific nicotinic acetylcholine receptor

“…Accordingly, in previous studies we have attempted to determine the major site of action of nAChR antagonists in promoting MN survival (Caldero et al, 1998;Oppenheim et al, 2000Oppenheim et al, , 2003 by, for example, using agents that act selectively at neuronal versus muscle nAChRs. Although the evidence from these studies is consistent with the selective role of muscle nAChRs in MN survival, reports of the presence of muscle-type nAChRs on chick embryo neurons (Pugh et al, 1995;Keiger et al, 2003) and of neuronal-type nAChRs in chick muscle Romano et al, 1997) have cast some doubt on whether agents such as curare and a-BTX rescue MNs by a peripheral versus CNS site of action (e.g., Hory-Lee and Frank, 1995;Usiak and Landmesser, 1999). For this reason, we have attempted to resolve this issue by the use of a novel cone snail toxin that is highly selective in binding to and antagonizing the embryonic/fetal form of muscle nAChRs (Teichert et al, 2004(Teichert et al, , 2005(Teichert et al, , 2006.…”

“…For example, we have previously shown in the chick embryo that drugs and toxins that selectively target muscle nAChRs results in the common phenotype described earlier, whereas agents that target neuronal nAChRs are ineffective (Oppenheim et al, 2000). Although these data are more consistent with a peripheral site of action for generating the common paralytic phenotype, because mRNAs for the adult a1 nAChR have been found in chick embryo neurons (Pugh et al, 1995;Keiger et al, 2003) and because mRNAs for neuronal nAChRs are transiently expressed in chick embryo muscle Romano et al, 1997;Keiger et al, 2003), we cannot entirely exclude the possibility of either a central site of action on adult muscle-type nAChRs or of a role for neuronal type nAChRs in muscle following treatment with most of the reagents used in previous studies to generate the paralytic phenotype. For example, the conotoxins EIVA, MI, and GI used in our previous study (Oppenheim et al, 2000) do not discriminate between adult versus fetal muscle-type nAChRs (Johnson et al, 1995;Jacobsen et al, 1997;Teichert et al, 2005).…”

Characterization of Rhythmic Ca2+Transients in Early Embryonic Chick Motoneurons: Ca2+Sources and Effects of Altered Activation of Transmitter Receptors

“…4A). The α7 subunit containing nicotinic receptors have a high permeability to Ca 2+ (Bertrand et al, 1993; Seguela et al, 1993) and are present in the lateral motor column (LMC) of embryonic chick spinal cords (Keiger et al, 2003). In addition, nicotinic signaling via these receptors regulates different aspects of neural development (Liu et al, 2006) including ACh mediated growth cone retraction of cultured motoneurons (Pugh and Berg, 1994).…”

Functional genomics of the nicotinic acetylcholine receptor gene family of the nematode, Caenorhabditis elegans

“…However, several discoveries, such as observations that certain neuronal nAChRs are expressed in developing muscle, (47) and the demonstration that the muscle a1 subunit is expressed in the developing central nervous system, (48) raise the possibility that more complex patterns of expression might be found. The green fluorescent protein (GFP), isolated from the jellyfish, Aequorea victoria, is an invaluable tool in studying gene expression in C. elegans.…”