Requirement of neuronal connexin36 in pathways mediating presynaptic inhibition of primary afferents in functionally mature mouse spinal cord

Abstract

Key points• Reflexes evoked by sensory information from muscles and skin play an important role in controlling muscle activity during movement.• The strength of these reflexes is regulated in part by presynaptic inhibition, a process controlling the release of chemical transmitters from sensory fibres terminating on spinal neurones.• Our study is the first to show that electrical synapses among spinal neurones in young animals are essential for normal operation of processes that presynaptically regulate synaptic transmission between large diameter sensory fibres and spinal cord neurones.• Transgenic mice lacking connexin36, a protein that mediates electrical communication via gap junctions between neurones, suffer a severe impairment of presynaptic inhibition and a similar impairment can be produced in normal mice with drugs that disrupt gap junction function.• The wide distribution of connexin36 in the spinal cord suggests that neuronal gap junctions also play a role in other physiological processes.Abstract Electrical synapses formed by gap junctions containing connexin36 (Cx36) promote synchronous activity of interneurones in many regions of mammalian brain; however, there is limited information on the role of electrical synapses in spinal neuronal networks. Here we show that Cx36 is widely distributed in the spinal cord and is involved in mechanisms that govern presynaptic inhibition of primary afferent terminals. Electrophysiological recordings were made in spinal cord preparations from 8-to 11-day-old wild-type and Cx36 knockout mice. Several features associated with presynaptic inhibition evoked by conditioning stimulation of low threshold hindlimb afferents were substantially compromised in Cx36 knockout mice. Dorsal root potentials (DRPs) evoked by low intensity stimulation of sensory afferents were reduced in amplitude by 79% and in duration by 67% in Cx36 knockouts. DRPs were similarly affected in wild-types by bath application of gap junction blockers. Consistent with presynaptic inhibition of group Ia muscle spindle afferent terminals on motoneurones described in adult cats, conditioning stimulation of an adjacent dorsal root evoked a long duration inhibition of monosynaptic reflexes recorded from the ventral root in wild-type mice, and this inhibition was antagonized by bicuculline. The same conditioning stimulation failed to inhibit monosynaptic reflexes in Cx36 knockout mice. Immunofluorescence labelling for Cx36 was found throughout the dorsal and ventral horns of the spinal cord of juvenile mice and persisted in mature animals. In deep dorsal horn laminae, where interneurones involved in presynaptic inhibition of large diameter muscle afferents are located, cells were extensively dye-coupled following intracellular neurobiotin injection. presynaptic inhibition, including the regulation of transmission from Ia muscle spindle afferents. In addition to a role in presynaptic inhibition in juvenile animals, the persistence of Cx36 expression among spinal neuronal populations in the adult mouse sugge...

Synaptic inhibition and excitation estimated via the time constant of membrane potential fluctuations

“…6D). The weak effects of spikes are likely due to the intense synaptic input present in this particular network (Berg et al 2008) causing the high-conductance state in the neuron and reducing the impact of intrinsic conductance (Berg and Hounsgaard 2009). We suggest that the ohmic estimate is a suitable comparison to evaluation of the ACF estimate.…”

“…The push-pull input is relatively easy to distinguish from the balanced E/I input, since the G tot is relatively constant in time in the push-pull. In the balanced input, G tot increases considerably compared with the degree of depolarization (Berg and Hounsgaard 2009). In balanced E/I, one would see a weak increase in the spiking during depolarization, whereas the conductance increase severalfold compared with G L .…”

“…For this reason, gap junctions represent a source of error in estimating the synaptic conductance. A way to verify gap-junction coupling is to apply a tracer Neurobiotin (Vector Laboratories, Burlingame, CA) in the electrode pipette, since it crosses most types of gap-junctions, and then perform a histological examination (Bautista et al 2012;Berkowitz et al 2006). Similarly, immunofluorescent labeling with Lucifer yellow for instance can identify the connexins constituting the gap junctions.…”

Connexin and pannexin signaling pathways, an architectural blueprint for CNS physiology and pathology?

“…In the mature brain, neuronal GJIC becomes more restricted and constitutes a distinct phenotypic feature of gamma-aminobutyric acid (GABA)ergic interneurons [32,105,108,109]. These electrical synapses are widely distributed over diverse regions of the mammalian brain, including the neocortex, hippocampus, thalamus, the inferior olive, the cerebellum as well as in the retina and the spinal cord [22,32,110,111]. Due to its widespread and dominant neuronal expression, Cx36 is considered to be the main molecular substrate of these synapses [110,112,113].…”

“…These electrical synapses are widely distributed over diverse regions of the mammalian brain, including the neocortex, hippocampus, thalamus, the inferior olive, the cerebellum as well as in the retina and the spinal cord [22,32,110,111]. Due to its widespread and dominant neuronal expression, Cx36 is considered to be the main molecular substrate of these synapses [110,112,113]. As GJCs play a pivotal role in the synchronization of electrical activity, they are also likely to facilitate the occurrence of epileptic seizures.…”