Spinal administration of lipoxygenase inhibitors suppresses behavioural and neurochemical manifestations of naloxone-precipitated opioid withdrawal

Abstract

1. This study investigated the role of spinal lipoxygenase (LOX) products in the induction and expression of opioid physical dependence using behavioural assessment of withdrawal and immunostaining for CGRP and Fos protein expression in the spinal cord. 2. Administration of escalating doses (5-50 mg kg-1; i.p.) of morphine for 5 days markedly elevated CGRP-like immunoreactivity in the dorsal horn of the rat spinal cord. Naloxone (2 mg kg-1; i.p.) challenge precipitated a robust withdrawal syndrome that depleted CGRP-like immunoreactivity and increased the number of Fos-like immunoreactive neurons in the dorsal horn. 3. Intrathecal administration of NDGA (10, 20 microg), a nonselective LOX inhibitor, AA-861 (1.5, 3 microg), a 5-LOX selective inhibitor, or baicalein (1.4, 2.8 microg), a 12-LOX selective inhibitor, concurrently with systemic morphine for 5 days or as a single injection immediately preceding naloxone challenge, blocked the depletion of CGRP-like immunoreactivity, prevented increase in the number of Fos-like immunoreactive neurons in the dorsal horn, and significantly attenuated the morphine withdrawal syndrome. 4. The results of this study suggest that activity of LOX products, at the spinal level, contributes to the expression of opioid physical dependence, and that this activity may be expressed through increased sensory neuropeptide release.

Figure 1

Effect of acute intrathecal pretreatment with LOX inhibitors on naloxone-induced morphine withdrawal. Animals were administered systemic morphine for 5 days, and given a single intrathecal drug injection 30 min prior to a naloxone challenge. The data are expressed as mean±s.e.m. Asterisks represent significant difference from morphine-treated animals challenged with naloxone: ^**P<0.01.

Figure 2

Effect of chronic intrathecal treatment with LOX inhibitors on naloxone-induced morphine withdrawal. LOX inhibitors were administered intrathecally in combination with systemic morphine for 5 days. At 3 h following the final dose on day 5, withdrawal was induced by naloxone (i.p.). The data are expressed as mean±s.e.m. Asterisks represent significant difference from morphine-treated animals challenged with naloxone: ^*P<0.05; ^**P<0.01.

Figure 3

Photomicrographs of CGRP-like immunoreactive axons in the dorsal horn of lumbar spinal cords in naive rats (a), following daily systemic injections of morphine for 5 days (b), and 1 h after naloxone-induced withdrawal (c). Withdrawal-associated depletion in CGRP-like immunoreactivity in the superficial laminae of the dorsal horn was attenuated by acute intrathecal treatment with NDGA (20 g) (d), AA-861 (3 g) (e), or baicalein (2.8 g) (f). This effect was also seen following chronic intrathecal administration with NDGA (20 g) (g), AA-861 (3 g) (h), or baicalein (2.8 g) (i). Scale bar 100 μm.

Figure 4

Effect of intrathecal treatment with LOX inhibitors on the mean optical density of CGRP-like immunoreactive axons in the lumbar region of the rat spinal dorsal horn. Asterisks represent significant difference from morphine-treated animals challenged with naloxone: ^**P<0.01.

Figure 5

Photomicrographs of Fos-like immunoreactive neurons in the dorsal horn of lumbar spinal cords in naive rats (a), following daily systemic injections of morphine for 5 days (b), and 1 h after naloxone-induced withdrawal (c). Induction of Fos during naloxone challenge was markedly suppressed by either acute or chronic intrathecal treatment with NDGA (20 g) (d; g), AA-861 (3 g) (e; h), or baicalein (2.8 g) (f; i). Scale bar 100 μm.

Figure 6

Effect of intrathecal administration of LOX inhibitors on the number of Fos-like immunoreactive neurons in the superficial and deeper laminae of the dorsal horn in lumbar spinal cords of rats. Asterisks represent significant difference from morphine-treated animals challenged with naloxone: ^*P<0.05; ^**P<0.01.