The first part of my thesis involves an investigation into mechanisms underlying the presynaptic regulation of transmitter release from myelinated hindlimb sensory afferents in rodents. The central hypothesis is that in addition to chemical transmission in spinal neuronal networks, electrical synapses formed by connexins are critically involved in presynaptic inhibition of large diameter sensory afferents. Subsequent sections of the thesis present a detailed examination of the distribution of connexins in the rodent spinal cord with a particular emphasis on the neuronal connexin, Cx36. Connexin36 (Cx36) is widely believed to be the protein forming the neuronal gap junctions that create electrical synapses between mammalian neurons in many areas of the central nervous system (Condorelli et al 1998). The first part of thesis concerns a previously unknown role of neuronal connexins in interneurone pathways involved in presynaptic control of synaptic transmission in the lumbar spinal cord of rodents. As far as we are aware, the idea that electrical contacts between spinal neurons contribute to spinal presynaptic inhibition is a novel hypothesis. Evidence will be presented: 1) that Cx36 is present in regions of the spinal cord containing interneurons involved in presynaptic inhibition, 2) that the lack of Cx36 in Cx36-/- knockouts mice results in a severe impairment of presynaptic inhibition, and 3) that blocking gap junctions pharmacologically in wild type mice impairs presynaptic inhibition. The exploration of this hypothesis will involve a combination of electrophysiological and immunohistochemical approaches in juvenile wild-type and knockout mice lacking Cx36, as well as immunohistochemical observations in adult rodents. This first section of the thesis begins with the development of a preparation in which several measures of presynaptic inhibition described in the in vivo adult cat preparation can be examined in vitro in young mice. The following sections of the thesis describe the distribution and features of Cx36 on neurons in mice and rats of different ages in four parts. The first will show that Cx36 is the only connexin associated with spinal neurons and refutes claims in the literature about the existence of a variety of connexions on spinal neurons. The second part will show that while gap junctions between some spinal neurons are only a transient developmental phenomenon, they persist in abundance in adult animals. The third part will present evidence of a previously unsuspected III association of Cx36 gap junctions at the chemical synapse between muscle afferent fibres and motoneurons. Specifically, an association between Cx36 and the glutamate transporter used in primary afferents, Vglut1 will be described. To our knowledge these results are the first to suggest the existence of mixed (electrical and chemical) synapses between primary afferents and motoneurons in the mature mammalian spinal cord. The final part of the thesis will describe the presence of Cx36 gap junctions on adult sacral motoneurons involved in control of sexual, urinary and defecation functions in the rodent.

The first part of my thesis involves an investigation into mechanisms underlying the presynaptic regulation of transmitter release from myelinated hindlimb sensory afferents in rodents. The central hypothesis is that in addition to chemical transmission in spinal neuronal networks, electrical synapses formed by connexins are critically involved in presynaptic inhibition of large diameter sensory afferents. Subsequent sections of the thesis present a detailed examination of the distribution of connexins in the rodent spinal cord with a particular emphasis on the neuronal connexin, Cx36. Connexin36 (Cx36) is widely believed to be the protein forming the neuronal gap junctions that create electrical synapses between mammalian neurons in many areas of the central nervous system (Condorelli et al 1998). The first part of thesis concerns a previously unknown role of neuronal connexins in interneurone pathways involved in presynaptic control of synaptic transmission in the lumbar spinal cord of rodents. As far as we are aware, the idea that electrical contacts between spinal neurones contribute to spinal presynaptic inhibition is a novel hypothesis. Evidence will be presented: 1) that Cx36 is present in regions of the spinal cord containing interneurones involved in presynaptic inhibition, 2) that the lack of Cx36 in Cx36-/- knockouts mice results in a severe impairment of presynaptic inhibition, and 3) that blocking gap junctions pharmacologically in wild type mice impairs presynaptic inhibition. The exploration of this hypothesis will involve a combination of electrophysiological and immunohistochemical approaches in juvenile wild-type and knockout mice lacking Cx36, as well as immunohistochemical observations in adult rodents. This first section of the thesis begins with the development of a preparation in which several measures of presynaptic inhibition described in the in vivo adult cat preparation can be examined in vitro in young mice. The following sections of the thesis describe the distribution and features of Cx36 on neurones in mice and rats of different ages in four parts. The first will show that Cx36 is the only connexin associated with spinal neurons and refutes claims in the literature about the existence of a variety of connexions on spinal neurons. The second part will show that while gap junctions between some spinal neurons are only a transient developmental phenomenon, they persist in abundance in adult animals. The third part will present evidence of a previously unsuspected III association of Cx36 gap junctions at the chemical synapse between muscle afferent fibres and motoneurones. Specifically, an association between Cx36 and the glutamate transporter used in primary afferents, Vglut1 will be described. To our knowledge these results are the first to suggest the existence of mixed (electrical and chemical) synapses between primary afferents and motoneurones in the mature mammalian spinal cord. The final part of the thesis will describe the presence of Cx36 gap junctions on adult sacral motoneurones involved in control of sexual, urinary and defecation functions in the rodent.