Re‐evaluation of connexins associated with motoneurons in rodent spinal cord, sexually dimorphic motor nuclei and trigeminal motor nucleus

Abstract

Electrical synapses formed by neuronal gap junctions composed of connexin36 (Cx36) are a common feature in mammalian brain circuitry, but less is known about their deployment in spinal cord. It has been reported based on connexin mRNA and/or protein detection that developing and/or mature motoneurons express a variety of connexins, including Cx26, Cx32, Cx36 and Cx43 in trigeminal motoneurons, Cx36, Cx37, Cx40, Cx43 and Cx45 in spinal motoneurons, and Cx32 in sexually dimorphic motoneurons. We re-examined the localization of these connexins during postnatal development and in adult rat and mouse using immunofluorescence labelling for each connexin. We found Cx26 in association only with leptomeninges in the trigeminal motor nucleus, Cx32 only with oligodendrocytes and myelinated fibers among motoneurons in this nucleus and in the spinal cord, and Cx37, Cx40 and Cx45 only with blood vessels in ventral horn of spinal cord, including those among motoneurons. By freeze-fracture replica immunolabelling (FRIL, >100 astrocyte gap junctions but no neuronal gap junctions were found based on immunogold labeling for Cx43, whereas 16 neuronal gap junctions at P4, P7 and P18 were detected based on Cx36 labelling. Punctate labelling for Cx36 was localized to the somatic and dendritic surfaces of peripherin-positive motoneurons in the trigeminal motor nucleus, motoneurons throughout the spinal cord, and sexually dimorphic motoneurons at lower lumbar levels. In studies of electrical synapses and electrical transmission between developing and between adult motoneurons, our results serve to focus attention on mediation of this transmission by gap junctions composed of Cx36.

Connexin36 identified at morphologically mixed chemical/electrical synapses on trigeminal motoneurons and at primary afferent terminals on spinal cord neurons in adult mouse and rat

“…2, if these neurons were subject to full 3D reconstruction. It is of note that while spinal neurons have been considered to express other connexins in addition to Cx36 (Chang et al, 1999), the above accordance between our results and earlier ultrastructural work further supports our observations reported elsewhere that neuronal gap junctions in adult spinal cord are formed primarily by Cx36 (Bautista et al 2013a). …”

“…Analysis of connexin mRNA and/or protein expression has suggested that multiple connexins are expressed in spinal and trigeminal motoneurons, including Cx26, Cx32, Cx36, Cx37, Cx40, Cx43 and Cx45 (Chang et al, 1999; Chang and Balice-Gordon, 2000; Chang et al, 2000; Personius and Balice-Gordon, 2000, 2001; Personius et al, 2001, 2007; Micevych and Abelson, 1991; Matsumoto et al, 1991,1992), raising the possibility that each of these connexins could form functional gap junctions between motoneurons. However, our recent analysis of connexin expression among trigeminal and spinal cord motoneurons in postnatal and adult rodent, using antibodies specific for each connexin, found labelling of only Cx36 in these neurons (Bautista et al, 2013a,b). Our results suggests that Cx36 is the major connexin mediating the electrical and dye-coupling observed between motoneurons at early postnatal ages.…”

“…Recently, we reported the expression of Cx36 in ventral and intermediate regions of the lumbar spinal cord of adult mice, and the loss of sensory-evoked presynaptic inhibition in mice lacking Cx36 (Bautista et al, 2012). The present study was motivated in part by our detection of abundant labelling of Cx36 in most spinal cord laminae, including extensive immunolabelling for Cx36 among motoneurons in lamina IX in adult rat and mouse (Bautista et al, 2012; Bautista et al, 2013a,b). The persistence of Cx36-puncta in adult spinal cord, particularly those associated with motoneurons, and the presence of ultrastructurally-identified gap junctions associated with some populations of motoneurons in adult rodent spinal cord (Matsumoto et al, 1988,1989; van der Want et al, 1998; Rash et al, 1996) appears to be at odds with the developmental loss of electrical coupling and dye-coupling between motoneurons (Fulton et al, 1980; Walton and Navarette, 1991; Chang et al, 1999).…”

Do gap junctions regulate synchrony in the Parkinsonian basal ganglia?

“…Our Cx36 antibody has been tested in Cx36-knockout mice [Bautista et al, 2014]. Only very slight background staining was present in this negative control (Bautista et al [2014], Fig. S1, supporting information).…”

“…However, we tried to minimize the effects of unblinded sampling by selecting tissue areas solely based on the DAPI signal. Our Cx36 antibody has been tested in Cx36-knockout mice [Bautista et al, 2014]. Only very slight background staining was present in this negative control (Bautista et al [2014], Fig.…”