Terriann Crisp

Analgesic effects of serotonin and receptor-selective serotonin agonists in the rat spinal cord.

1. Serotonin (5-HT) and selective 5-HT receptor agonists were administered intrathecally (i.t.) in rats, and the antinociceptive efficacy of these agents was assessed on the tail-flick and hot plate tests. 2. The 5-HT receptor agonists examined in this study included the 5-HT1A agonist 8-hydroxy-N,N-dipropyl-2-aminotetralin (8-OH-DPAT), the 5-HT1B agonist m-trifluoromethylphenylpiperazine (TFMPP), the 5-HT2 agonist 1-(2,5-dimethoxy-4-iodophenyl)-2-aminopropane (DOI) and the 5-HT3 agonist phenylbiguanide (PBG). 3. None of these agents produced significant elevations in tail-flick latency (TFL) at doses which produced elevations in hot plate latency (HPL). 4. In contrast, the i.t. dose of 5-HT which elevated TFL also produced analgesia on the hot plate test. 5. Serotonin-induced elevations in TFL were reversed by pindolol, ritanserin and ICS 205-930, suggesting that 5-HT interacts with more than one 5-HT site in the spinal cord to produce analgesia on the tail-flick test. 6. The finding that ritanserin reversed 5-HT-induced elevations in HPL suggests that the 5-HT2 site is primarily responsible for mediating the spinal antinociceptive effects of 5-HT on the hot plate test.

The effects of aging on spinal neurochemistry in the rat

The purpose of this study was to investigate how the aging process alters the basal levels of serotonin, norepinephrine, dopamine, and their respective metabolites in the spinal cord using high-performance liquid chromatography and electrochemical detection. Young, mature and aged male Fischer 344 rats (5-6, 15-16, and 25-26 months old, respectively) were used in all experiments. Dorsal and ventral halves of the cervical, thoracic, lumbar and sacral vertebral sections of the rat spinal cord were analyzed. The results indicate that as chronological age increases, local spinal levels of serotonin and norepinephrine decrease. These findings are discussed in terms of how age-related changes in the endogenous levels of the biogenic amines may alter the perception of pain in the elderly.

Microglial proliferation in the spinal cord of aged rats with a sciatic nerve injury

Nerve injury may lead to chronic neuropathic pain syndromes. We determined whether the extent of central nervous system microglial activation that accompanies nerve injury is age dependent and correlated with behavioral manifestations of pain. We used the Bennett and Xie sciatic nerve chronic constriction injury model (Bennett, G.J., Xie, Y.-K., A peripheral mononeuropathy in rat that produces disorders of pain sensation like those seen in man, Pain, 33 (1998) 87-107) to induce neuropathic pain in three age cohorts of Fischer 344 FBNF1 hybrid rats (4-6, 14-16, and 24-26 months). Rats were assessed for thermal sensitivity (hyperalgesia) of their hind paws pre-injury (day 0) and up to 35 days post injury. On various days post injury, the L4-L5 levels of their spinal cords were reacted for localization of an antibody to OX-42, a marker for microlgia. OX-42 immunoreactivity (ir) was quantified by use of a Bioquant density analysis system. OX-42 ir was heavy in areas of sciatic nerve primary afferent terminations and in the motor columns of its neurons. Aging increases OX-42 ir in the absence of injury. After injury, OX-42 ir increased further, but the increases over control levels decreased with age. Ligation-induced analgesia and hyperalgesia were both correlated with the increases in OX-42 ir, regardless of age.

Gender and the behavioral manifestations of neuropathic pain.

A model of peripheral nerve injury was used to study gender differences in the development and progression of chronic constriction injury (CCI)-induced hyperalgesia and allodynia in male and female Fischer 344 FBNF1 hybrid rats. Rats were randomly assigned to one of the following treatment groups: (1) gonadally intact unligated males (male); (2) gonadally intact ligated males (male (CCI)); (3) castrated ligated males (male (CAS/CCI)); (4) gonadally intact unligated females (female); (5) gonadally intact ligated females (female (CCI)); and (6) ovariectomized ligated females (female (OVX/CCI)). A plantar analgesia meter and calibrated von Frey pressure filaments were used as the analgesiometric assays. In the absence of nerve injury, gonadally intact males responded significantly faster than females to a thermal nociceptive stimulus. The onset of the behavioral manifestations of unilateral ligation of the sciatic nerve did not differ as a function of sex or hormonal status (e.g., gonadally intact and gonadectomized male and female rats developed thermal hyperalgesia within 14 days post-CCI). Paw withdrawal latency (PWL) values of gonadally intact males returned to baseline control values after postligation day 14, whereas gonadally intact females, ovariectomized females and castrated males continued to elicit robust thermal hyperalgesic symptoms throughout the 35-day duration of the experiment. Allodynic responses to peripheral nerve injury were less variable across genders. These data suggest that the mechanisms underlying chronic nociceptive processing differ as a function of gender and gonadal hormone status.

Effects of aging on spinal opioid-induced antinociception

Initial experiments were conducted to determine whether or not the aging process alters the ability of young, mature, or aged male Fischer 344 rats (5- to 6-, 15- to 16-, and 25- to 26-months-old, respectively) to respond to thermal nociceptive stimuli. Using the tail-flick analgesiometric assay, 25- to 26-month-old rats responded significantly faster to the heat source than 15- to 16-month-old animals, but no significant differences were noted between the 5- to 6-month-old and aged rats. Another series of investigations compared the effects of aging on the spinal antinociceptive properties of the mu opioid agonist [D-Ala2,N-methyl-Phe4,Gly5-ol] enkephalin (DAMPGO) and the delta agonist [D-Pen2,D-Pen5] enkephalin (DPDPE). In these studies, young, mature, and aged rats were injected intrathecally (IT) with different doses of DAMPGO or DPDPE, and opioid-induced antinociception was tested on the tail-flick test. All three age groups responded to IT DAMPGO in a dose-dependent manner but, for the most part, higher spinal doses were required to produce significant elevations in tail-flick latency in the aged cohort of rats. The spinal analgesic effects of DPDPE also declined with advanced age. The aging process apparently alters the pain-inhibitory function of mu and delta opioid receptors in the rat spinal cord.

Serotonin contributes to the spinal antinociceptive effects of morphine

This study was designed to determine if morphine administered intrathecally (IT) interacts with serotonergic or noradrenergic nerve terminals in the spinal cord to produce analgesia on the spinally mediated tail-flick test. Male Sprague-Dawley rats were fitted with IT catheters. One week later, animals were spinally pretreated with receptor antagonists selective for opioid, serotonin or alpha-adrenoceptors, and the ability of these agents to alter spinal morphine-induced antinociception was assessed. Morphine dose-dependently elevated tail-flick latency in a naltrexone-reversible manner. The serotonin receptor antagonists spiroxatrine (5-HT1A), pindolol (5-HT1B), ritanserin (5-HT2) and ICS 205-930 (5-HT3) attenuated the spinal analgesic effects of morphine. In contrast, the alpha 1 and alpha 2-adrenoceptor antagonists prazosin and yohimbine, respectively, did not alter morphine-induced elevations in tail-flick latency. These data substantiate earlier reports that spinal morphine-induced antinociception relies on an opioid receptor-mediated component in addition to a local serotonergic component. The finding that the alpha-adrenoceptor antagonists did not alter the antinociceptive effects of IT morphine suggests that spinal norepinephrine does not contribute to the analgesic effects of the opiate.

A single restraint stress exposure potentiates analgesia induced by intrathecally administered DAGO

In rats, restraint exposure potentiates the magnitude and duration of analgesia following both the peripheral and intracerebroventricular administration of several opioid agonists as compared to non-stressed controls. It has been suggested that the site of action whereby restraint leads to potentiated opioid analgesia is located supraspinally. However, the possible contribution of spinal analgesic mechanisms also warrants investigation. Thus, the purpose of the present study was two-fold: (1) to determine whether a single exposure to restraint stress would result in the dose-dependent potentiation of analgesia following the intrathecal (i.t.) administration of the mu (mu)-receptor selective opioid agonist [D-Ala2,N-Me-Phe4,Gly5-ol]enkephalin (DAGO) and (2) to quantify the degree of analgesia in restrained vs. non-restrained rats using the tail-flick and hot-plate analgesic assays. Using rats implanted with chronic i.t. cannula, dose- and time-course curves were observed following the i.t. administration of DAGO. The results demonstrate that both the duration and magnitude of analgesia was significantly potentiated in restrained rats compared to non-restrained controls. Restraint-treated rats receiving 0.15-0.6 micrograms of DAGO i.t. showed 1.3-1.5-fold potentiation of analgesia in the tail-flick assay and a 2.3-5.6-fold potentiation using the hot-plate assay. Restraint immobilization potentiated the magnitude and duration of DAGO-induced analgesia administered by the i.t. route as measured by the tail-flick and hot-plate assays. These data suggest that spinal analgesic mechanisms significantly contribute to the enhanced analgesic potency of opioids in subjects exposed to restraint stress.

Spinal β-endorphin analgesia involves an interaction with local monoaminergic systems

beta-Endorphin administered intrathecally (i.t.) in rats produced a dose-dependent elevation in tail-flick latency. Naltrexone administered i.t. as a pretreatment reversed the spinal antinociceptive action of beta-endorphin, suggesting that the opioid interacts directly with spinal opiate receptors. Spinal administration of the alpha 1-adrenoceptor antagonist WB-4101 failed to alter the analgesic effects of the opioid, whereas the alpha 2-adrenoceptor antagonist yohimbine completely blocked beta-endorphin-induced elevations in tail-flick latency. Thus, there is an apparent specificity for the alpha 2-adrenoceptor to mediate the spinal action of beta-endorphin. The 5-HT1 and 5-HT3 receptor antagonists (spiroxatrine and ICS 205-930, respectively) also reversed the analgesic effects of the opioid, while the 5-HT2 receptor antagonist ritanserin only partially blocked beta-endorphin-induced elevations in tail-flick latency. The present results suggest that beta-endorphin produces analgesia at the spinal level via an opiate receptor-mediated interaction with spinal monoaminergic nerve terminals.

Effect of aging on the substance P receptor, NK-1, in the spinal cord of rats with peripheral nerve injury.

Substance P (SP) levels in the spinal cords of very old rats are less than the levels in younger rats (Bergman et al., 1996). After injury to a peripheral nerve in young rats, immunoreactivity (ir) to the SP receptor, NK-1 (neurokinin-1), increases in the spinal cord ipsilateral to the injury and the increases are correlated with the development of thermal hyperalgesia (Goff et al., 1998). Thus we postulated that aged rats might display an increased sensitivity to thermal stimulation before peripheral nerve injury and that they might respond differently to injury than do younger rats. To test this hypothesis, we used the Bennett and Xie model (1988) of chronic constriction injury (CCI) to the sciatic nerve to induce a neuropathic pain condition. We investigated the effect of age on changes in NK-1 ir in superficial layers of the dorsal horn and on numbers of NK ir cells in deeper laminae at the L4-L5 levels of the spinal cord after CCI. NK-1 receptors were tagged immunohistochemically and their distribution quantified by use of computer-assisted image analysis. NK-1 ir changes were related to alterations in thermal and tactile sensitivity that developed after CCI in young, mature and aged (4-6, 14-16, and 24-26 months) Fischer F344 BNF1 hybrid rats. No differences in thermal or tactile sensitivity of young and aged rats were seen in the absence of nerve injury. After injury, aged rats developed thermal hyperalgesia and tactile allodynia more slowly than did the younger rats. NK-1 receptor ir and numbers of NK-1 ir cells in the dorsal horn increased with time post-injury in all three groups. NK-1 ir increases were correlated with the development of thermal hyperalgesia in those rats that displayed hyperalgesia. However, some rats developed an increased threshold to thermal stimuli (analgesia) and that also was correlated with increases in NK-1 ir. Thus NK-1 ir extent, while correlated with thermal sensitivity in the absence of injury, is not a specific marker for disturbances in one particular sensory modality; rather it increases with peripheral nerve injury per se.Substance P (SP) levels in the spinal cords of very old rats are less than the levels in younger rats (Bergman et al., 1996). After injury to a peripheral nerve in young rats, immunoreactivity (ir) to the SP receptor, NK–1 (neurokinin-1), increases in the spinal cord ipsilateral to the injury and the increases are correlated with the development of thermal hyperalgesia (Goff et al., 1998). Thus we postulated that aged rats might display an increased sensitivity to thermal stimulation before peripheral nerve injury and that they might respond differently to injury than do younger rats. To test this hypothesis, we used the Bennett and Xie model (1988) of chronic constriction injury (CCI) to the sciatic nerve to induce a neuropathic pain condition. We investigated the effect of age on changes in NK-1 ir in superficial layers of the dorsal horn and on numbers of NK ir cells in deeper laminae at the L4-L5 levels of the spinal cord after CCI. NK-1 receptors were tagged immunohistochemically and their distribution quantified by use of computer-assisted image analysis. NK-1 ir changes were related to alterations in thermal and tactile sensitivity that developed after CCI in young, mature and aged (4-6, 14-16, and 24-26 months) Fischer F344 BNF1 hybrid rats. No differences in thermal or tactile sensitivity of young and aged rats were seen in the absence of nerve injury. After injury, aged rats developed thermal hyperalgesia and tactile allodynia more slowly than did the younger rats. NK-1 receptor ir and numbers of NK-1 ir cells in the dorsal horn increased with time post-injury in all three groups. NK-1 ir increases were correlated with the development of thermal hyperalgesia in those rats that displayed hyperalgesia. However, some rats developed an increased threshold to thermal stimuli (analgesia) and that also was correlated with increases in NK-1 ir. Thus NK-1 ir extent, while correlated with thermal sensitivity in the absence of injury, is not a specific marker for disturbances in one particular sensory modality; rather it increases with peripheral nerve injury per se.

Age-related changes in the spinal antinociceptive effects of DAGO, DPDPE and β-endorphin in the rat

These studies were designed to investigate how the aging process alters the spinal antinociceptive efficacy of mu (μ), delta (δ) and epsilon (e) opioid receptor agonists administered intrathecally (i.t.) in rats. Various doses of the μ agonist DAGO, the δ agonist DPDPE or the putative e agonist β-endorphin were injected i.t. in young (5–6-month-old), mature (15–16-month-old) and aged (25–26-month-old) Fischer 344 rats. Antinociception was measured using the rat tail-flick analgesiometric assay. The data demonstrated a decline in spinal opioid-induced antinociception as a function of age. For instance, the i.t. dose of DPDPE or β-endorphin needed to produce antinociception in the 25–26-month-old rats was higher than that needed to elevate tail-flick latency in the young and mature animals. We also noted that the i.t. doses of the opioid agonists needed to produce ‘antinociception’ in the aged cohort were within a range of spinal doses that produced motor impairment. Apparently, the aging process alters the ability of opioid receptors to mediate antinociception. Perhaps an age-related decrease in the number and/or affinity of opioid receptor sites in the rat spinal cord accounts for these observations.