Riley, Ruth Clare

2018-05-14T10:15:43Z

2018-05-14T10:15:43Z

1996

Although much is now known of the cellular actions of exogenous opiate drugs and of endogenous opioid peptides, how these compounds modify interconnecting neuronal systems in the CNS is still poorly understood. The studies described in this thesis have addressed the latter by employing the antibody microprobe technique (Duggan A.W. and Hendry I.A., 1986, Neurosci. Letts. 68, 134-140) to investigate (a) the release of dynorphin A(l-8) in the spinal cord of the rat as inflammation develops in peripheral tissues, and (b) the release of cholecystokinin, a putative 'anti-opioid' neuropeptide, in the rat spinal cord following the administration of morphine.

(a) A dramatic increase in the spinal synthesis of prodynorphin derived peptides has been observed when inflammation develops in peripheral tissues. The functional significance of this increased synthesis is unclear and there have been no reports of the stimuli needed to produce dynorphin release in vivo nor of possible controls of such release. Microprobes bearing immobilised antibodies to the dynorphin A(l-8) derivative of prodynorphin, were inserted into the lumbar spinal cord of urethane anaesthetised normal rats and those with a peripheral inflammation, to determine whether dynorphins are being tonically released and how release is altered by manipulating the inflamed tissues. In the absence of any active peripheral stimulus the antibody microprobes detected minimal amounts of immunoreactive (ir)-dynorphin A(l-8) in two areas (lamina I and laminae IV-V) in the dorsal horn of the spinal cord of normal rats. With the development of unilateral ankle inflammation over 3 to 5 days, following subcutaneous injections of Freund's complete adjuvant, this was extended to the ventral horn of both sides of the spinal cord. Lateral compression of the ankles of the normal animals did not release ir-dynorphin A(l-8) during the period of stimulation, but this neuropeptide was detected in the ventral horn following the stimulus. By contrast, compression of inflamed ankles produced elevated levels of ir-dynorphin A( 1 -8) during the period of stimulus application at three major sites in the spinal grey matter. The largest peak was in the deep dorsal horn/ upper ventral horn (laminae VI -VII), with further sites of significant release in the mid dorsal horn (laminae II-V) and the lower ventral horn. These levels persisted for at least one hour after the period of stimulation. At a cellular level dynorphins reduce transmitter release from nerve terminals, and hence the observation that ir-dynorphin A( 1 -8) is released in the ventral and deep dorsal horn in addition to the superficial dorsal horn of the rat by manipulation of inflamed tissues, implies that wide-spread spinal inhibitory controls of spinal neuronal firing are evoked by such stimuli. This may ultimately affect the perception of pain, but release in the ventral horn suggests an involvement in reducing motor responses to peripheral noxious stimuli.

(b) Of all possible candidates at the spinal cord level, the 'anti-opioid' activity of cholecystokinin (CCK) has been well characterised. This peptide has also been proposed to play a role in the development of tolerance to but not dependence on opiate drugs. Although the hypothesis that stimulation of opioid receptors may trigger a progressive compensatory increase in the activity of CCK containing neurones at the spinal cord level has received some indirect support, this has not been thoroughly investigated. Conflicting data have been obtained from experiments which have examined the spinal cord content of CCK and spinal release of CCK following the administration of opioids. Microprobes bearing immobilised antibodies to CCK-8, were inserted into the lumbar spinal cord of urethane anaesthetised normal rats under both basal conditions and following acute opiate administration. In the absence of any active peripheral stimulus an extensive presence of ir-CCK was detected in normal (drug naive) rats with three main zones. The largest peak was in the mid to deep dorsal horn/ upper ventral horn (laminae III -VII), with further major sites in the superficial dorsal horn (lamina I-II) and the mid/ lower ventral horn. Morphine administered intravenously over two hours (total dose 25mg/ kg) failed to alter this basal presence of irCCK. Enhanced release however, was observed in areas of the ventral horn of rats treated acutely with morphine following lmg/ kg injections of naloxone. It is proposed that the occupation of opioid receptors by morphine triggers the increased synthesis of CCK and predicted that following the development of tolerance to morphine, enhanced CCK release will be observed in the absence of naloxone administration.

http://hdl.handle.net/1842/29964

The University of Edinburgh

Annexe Thesis Digitisation Project 2018 Block 18

Already catalogued

Studies of dynorphin and cholecystokinin release in the rat spinal cord: implications for opioid action

Thesis or Dissertation

Doctoral

PhD Doctor of Philosophy