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Tracing axons and axon collaterals of spinal neurons using intracellular injection of horseradish peroxidase.

Intracellular injection and subsequent histochemical localization of horseradish peroxidase have been used to stain the soma, dendrites, axons, and axon collaterals of spinalcervical tract neurons and unidentified dorsal horn neurons in the cat. This technique may be used in combination with the intracellular injection of Procion yellow to demonstrate by light microscopy connections between physiologically typed vertebrate neurons.

Animals↗

Attenuation of morphine tolerance after antisense oligonucleotide knock-down of spinal mGluR1.

1. Chronic systemic treatment of rats with morphine leads to the development of opioid tolerance. This study was designed to examine the effects of intrathecal (i.t.) infusion of a metabotropic glutamate receptor 1 (mGluR1) antisense oligonucleotide, concomitant with chronic morphine treatment, on the development of tolerance to morphine's antinociceptive effects. 2. All rats received chronic (6 day) s.c. administration of morphine to induce opioid tolerance. Additionally, rats were treated with either mGluR1 antisense (AS), missense (MIS) or artificial cerebrospinal fluid (ACSF) by i.t. infusion via chronically implanted i.t. catheters connected to osmotic mini-pumps. The effects of acute i.t. or s.c. morphine on tail-flick latencies were assessed prior to and following chronic s.c. morphine treatment for all chronic i.t. infusion groups. mGluR1 protein level in the spinal cord was determined by Western blot analysis for all treatments, assessing the efficiency of knock-down with AS treatment. 3. Acute i.t. morphine dose-dependently produced antinociception in the tail-flick test in naïve rats. Systemic morphine-treated rats administered i.t. ACSF or MIS developed tolerance to i.t. morphine. Chronic i.t. infusion with mGluR1 AS significantly reduced the development of tolerance to i.t. morphine. 4. In contrast to i.t. morphine, tolerance developed to the antinociceptive effects of s.c. morphine, in all i.t. infusion groups, including the mGluR1 AS group. 5. The spinal mGluR1 protein level was dramatically decreased after mGluR1 AS infusion when compared to control animals (naïve and ACSF-treated animals). 6. These findings suggest that the spinal mGluR1 is involved in the development of tolerance to the antinociceptive effects of morphine. Selective blockade of mGluR1 may be beneficial in preventing the development of opioid analgesic tolerance.

Analgesics, Opioid↗

Lumbosacral anterolateral spinal arteries and brief review of "accessory" longitudinal arteries of the spinal cord.

The lumbosacral anterolateral spinal arteries (LALSA) were studied in 14 injected lumbosacral spinal cords. Contrary to many previously published opinions, which claimed that virtually all of the formerly described "accessory" longitudinal arteries of the vasa corona, were too inconstant to be of significance, the LALSA in this series were reliably present bilaterally and averaged a length of 8.4 cm. They ran in the acute angles formed by the emergence of the fascicles of the lumbosacral anterior spinal nerve roots and in this position, served as the origin for most of the proximal radicular arteries that supply the cranial half of the motor roots of the cauda equina. A review of the literature and analysis of other accessory longitudinal spinal arteries revealed that two other pairs of accessory spinal vessels, the lateral cervical spinal arteries (LCSA) and the lateral spinal arterial axes (LSA), have a demonstrable functional role and were sufficiently constant, along with the LALSA, to warrant inclusion in detailed descriptions of human spinal cord vasculature.

Cadaver↗

Somatostatinergic neurons in the insular cortex project to the spinal cord: combined retrograde axonal transport and immunohistochemical study.

A double-labeling method combining immunohistochemistry and a retrograde tracer technique using biotin-horseradish peroxidase (B-HRP) was employed to identify a descending somatostatinergic fiber system from the insular cortex to the spinal cord. Injection of B-HRP into the spinal cord at cervical or lumbar levels resulted in the labeling of a number of neurons in the insular cortex. Simultaneous immunostaining revealed the existence of double-labeled neurons in the insular cortex. The result provides direct evidence for the presence of a descending somatostatinergic pathway from the insular cortex to lumbar levels of the spinal cord.

Animals↗

Stimulation of spinal delta-opioid receptors in mice selectively enhances the attenuation of delta-opioid receptor-mediated antinociception by antisense oligodeoxynucleotide.

Intrathecal (i.t.) pretreatment of male ICR mice with antisense oligodeoxynucleotide to delta-opioid receptor mRNA once a day for 1-3 days caused a time-dependent attenuation of i.t. administered [D-Ala2]deltorphin II-induced antinociception as measured by the tail-flick test. The attenuation of the antinociception induced by i.t. administered [D-Ala2]deltorphin II, a delta-opioid receptor agonist, was enhanced by i.t. pretreatment for 1 day with [D-Ala2]deltorphin II, but not [D-Ala2,N MePhe4,Gly(ol)5]enkephalin (DAMGO), a mu-opioid receptor agonist, or U50,488H, a kappa-opioid receptor agonist, given together with antisense oligodeoxynucleotide to delta-opioid receptor mRNA. The present results indicate that stimulation of spinal delta-opioid receptors by i.t. injection of [D-Ala2]deltorphin II selectively causes a loss of delta-opioid receptor-mediated antinociception in mice pretreated with antisense oligodeoxynucleotide to delta-opioid receptor mRNA.

Analgesics↗

Slow axonal transport is impaired by intrathecal 2,5-hexanedione.

Anterograde axonal transport was studied in a new model of hexacarbon neuropathy, in which neurofilament (NF)-containing giant axonal swellings are induced proximally in the spinal nerve roots of rats by intrathecal injection of 2,5-hexanedione (2,5-HD). Decreased transport velocity of the NF-containing slow component a (SCa) was demonstrated in 2,5-HD-treated animals, in contrast to studies demonstrating increased velocity of SCa in proximal parts of the axon in systemic 2,5-HD intoxication, which causes distal axonal swellings. Other components of anterograde transport were unaffected. In systemic 2,5-HD toxicity, velocity of NF transport increases in the proximal axon, but may decrease distally, where it is difficult to study. Decreased NF transport is likely to be responsible for the formation of axonal swellings, since they occur preterminally rather than at the axon terminal as would be expected if increased NF transport were the cause. Covalent modification of proteins provides a possible mechanism by which 2,5-HD affects axonal transport, and the effect may be facilitatory or inhibitory depending on the level and duration of exposure of the NF to the toxin.

Animals↗

Inhibitory effect of intrathecally administered nociceptin on the expression of Fos-like immunoreactivity in the rat formalin test.

Nociceptin, an opioid receptor-like 1 receptor agonist, has been reported to produce an analgesic effect in the rat formalin test. The authors examined the effect of 17 nmol of nociceptin on the expression of Fos-like immunoreactivity (Fos-LI) in the spinal dorsal horn induced by paw formalin injection and compared the effect of 17 nmol of nociceptin with that of equiantinociceptive dose (3 nmol) of morphine. Nociceptin and morphine were administered intrathecally 10 min before the formalin injection and the expression of Fos-LI was examined 2.5 h after the formalin injection. Both 17 nmol of nociceptin and 3 nmol of morphine suppressed the expression of Fos-LI in laminae I-II, but not in laminae III-V, to the same extent. These data suggest that intratheally administered nociceptin and morphine suppress the nociceptive input into the laminae I-II and produce antinociceptive effect.

Analgesics↗

Locomotor projections from the pedunculopontine nucleus to the spinal cord.

Following injections of retrograde tracers into the spinal cord enlargements, the mesencephalic locomotor region (MLR) was electrically stimulated to induce locomotion in decerebrate rats. MLR sites coincided with cholinergic pedunculopontine nucleus (PPN) neurons, which were selectively labelled using NADPH diaphorase histochemistry. Retrogradely labelled (spinal-projecting) PPN neurons were scattered among cholinergic PPN neurons. The absence of retrogradely labelled, NADPH diaphorase positive PPN neurons indicates that spinal enlargement projections are non-cholinergic.

Animals↗

Intrathecal sufentanil labor analgesia: the effects of adding morphine or epinephrine.

Intrathecal opioids can provide labor analgesia. We attempted to prolong the duration of intrathecal sufentanil analgesia by adding epinephrine or morphine. Forty-one healthy, term nulliparae with cervical dilation < 5 cm participated in this double-blind, randomized protocol. Using a combined spinal and epidural technique, we gave intrathecal injections of either sufentanil 10 micrograms, sufentanil 10 micrograms plus epinephrine 200 micrograms, or sufentanil 10 micrograms plus morphine 250 micrograms. At baseline and every 5 min for 30 min thereafter, we recorded arterial blood pressure and asked the patients to rate their pain, nausea, and pruritus on visual analog scales. The women continued to rate these variables every 30 min until they requested additional analgesia. They then received 10 mL of 0.25% bupivacaine via the epidural catheter, and rated the above variables every 30 min until they requested additional epidural drug injection. Both morphine and epinephrine prolonged the duration of sufentanil analgesia. Only morphine prolonged analgesia after the first dose of epidural bupivacaine. However, because women in the morphine group experienced significantly more side effects throughout the study period, we do not recommend intrathecal morphine for labor analgesia.

Adult↗

Role of gap junction in the expression of morphine-induced antinociception.

The present study was undertaken to investigate whether gap junctional communication could be involved in morphine-induced antinociceptive response using blockers of the gap junctional channel, carbenoxolone and Gap27. Intrathecal pretreatment with either carbenoxolone or Gap27 caused a dose-dependent attenuation of morphine-induced antinociception. Furthermore, the dose-response line for morphine-induced antinociception was shifted to the right by 2.53-fold following intrathecal treatment with carbenoxolone. These findings suggest that gap-junctional-dependent communication in the mouse spinal cord may play, at least in part, a role in the expression of morphine-induced antinociception.

Analgesics, Opioid↗

Injections of calcium ions into spinal motoneurones.

1. In cats under Dial anaesthesia, Ca(2+) was injected inside lumbosacral motoneurones, by passing currents between CaCl(2)- and KCl-containing barrels of compound micropipettes.2. There was a reduction in excitability and a fall in membrane resistance, both rapid in onset and quickly reversible.3. The minimum effective injection current was approximately 10 nA, and the effect reached a maximum with currents of approximately 30 nA. The mean slope of resistance change against injection current was -1.7%/nA (S.E. 0.35).4. The most common change in membrane potential was a hyperpolarization; but in nearly half the cases, there was no clear change or a small depolarization. A reversal level for the effect of Ca(2+) could be measured in five cells: on the average, it was 10 mV more negative than the resting potential.5. Observations on i.p.s.p.s showed that Ca(2+) probably does not alter g(Cl): it was concluded that the fall in membrane resistance caused by intracellular Ca(2+) is mainly due to an increase in g(K).6. These results confirm previous suggestions that a steep transmembrane gradient of Ca(2+) is essential for the maintenance of a low membrane conductivity, and that a rise in internal free Ca(2+) - whether due to influx or release from internal stores - may play an important role in regulating neuronal activity.

Action Potentials↗

Vaccinia virus complement control protein reduces inflammation and improves spinal cord integrity following spinal cord injury.

Traumatic spinal cord injury (SCI) claims approximately 10,000 new victims each year in the United States alone. The injury usually strikes those under the age of 30 years, often leading to a lifetime of pain, suffering, and disability. Therapeutic agents targeting spinal cord injury are sorely lacking, and therefore our laboratory endeavored to evaluate the potential therapeutic benefits of immediate post-injury administration of the vaccinia virus complement control protein (VCP). VCP is a multifunctional anti-inflammatory protein that can inhibit both pathways of complement activation and bind heparin. Utilizing a common animal model of contusion SCI, motor function recovery tests, and immunochemical stains, we evaluated the effects of VCP injected into spinal cord tissue following injury. Results demonstrate that VCP administration inhibits macrophage infiltration, reduces spinal cord destruction, and improves hind-limb function, establishing VCP as a strong candidate for further investigation in the treatment of SCI.

Animals↗

Systemic physostigmine increases the antinociceptive effect of spinal morphine.

In this study we evaluated the antinociceptive effect of concurrent intrathecal (i.t.) and subcutaneous (s.c.) administration of morphine and physostigmine, respectively. The experiments were performed on male Wistar rats. Intrathecal administration of morphine was performed through a catheter implanted in the subarachnoid space. The 'tail-immersion' test was used to measure animals' responses to evoked nociceptive stimuli. Interaction of drugs was analyzed using a dose addition model. Both i.t. (1-5 microg) administration of morphine and s.c. (50-250 microg/kg) administration of physostigmine increased the latencies of nociceptive responses in a dose-dependent manner. Two micrograms of i.t. morphine and 100 microg/kg of s.c. physostigmine demonstrated 31.6 +/- 10.6 and 34.2 +/- 11.4 percentage of maximal possible effect (%MPE), respectively. Simultaneous administration of 1 microg of i.t. morphine and 50 microg/kg of s.c. physostigmine produced a %MPE equal to 84.8 +/- 16.9. Thus, combined administration of 1 microg i.t. morphine and 50 microg/kg s.c. physostigmine resulted in a strong, highly significant antinociceptive effect. This effect was much higher than the effect expected if both drugs acted in an additive manner. Supra-additive interaction observed in this study might be a result of simultaneous activation of different neurotransmitter systems involved in nociceptive processing at the spinal as well as at the supraspinal level of the CNS.

Analgesics, Opioid↗

The spinal and peripheral roles of bradykinin and prostaglandins in nociceptive processing in the rat.

The effect of peripheral and intrathecal administration of the bradykinin B2 receptor antagonist HOE140 and the non-steroidal anti-inflammatory drug indomethacin were studied on the response of dorsal horn nociceptive neurones to formalin in the anaesthetized rat. Peripheral pretreatment with HOE140 reduced both phases of the formalin response whereas subcutaneous administration of indomethacin (5 mg/kg) reduced only the second phase. Pre-treatment with intrathecal HOE140 resulted in a dose-dependent reduction of the second phase whereas a high dose (500 micrograms) intrathecal indomethacin reduced both phases of the response. Bradykinin plays a role in peripheral nociception during the first and second phase of the response whereas the prostaglandins are only involved during the second phase. The converse is true centrally, the prostaglandins appear to be involved in both phases of the formalin response whereas bradykinin only plays a central role during the second phase of the response.

Animals↗