Effect of opiates on the release of cholecystokinin from in vitro hypothalamus and frontal cortex of Zucker lean (Fa/−) and obese (fa/fa) rats

Opiates, morphine and [d-Ala²-d-Leu⁵]-enkephalin (DADLE), inhibited the K⁺-stimulated release of cholecystokinin (CCK) from the hypothalamus of both Zucker obese (fa/fa) and lean (Fa/−) rats, in vitro. Morphine and DADLE did not inhibit the K⁺-stimulated release of CCK from frontal cortex from either strain. The opiates did not affect basal efflux of CCK and their effects were all blocked by equimolar concentrations of naloxone. These studies indicate a regional specificity for the effect of opiates on CCK release, and may provide evidence for a cellular mechanism by which endogenous opiates modulate feeding behavior.     

Antagonism by indomethacin of neurogenic hyperthermia produced by unilateral puncture of the anterior hypothalamic/preoptic region

1. In unanaesthetized rats, restrained at an ambient temperature of 24 degrees C, the anterior hypothalamic/preoptic (AH/PO) region was lesioned unilaterally by acute mechanical puncture.2. In control (no pre-treatment) rats, unilateral AH/PO puncture produced a neurogenic hyperthermia which began immediately, reached its peak magnitude (mean peak magnitude = +2.3 degrees C) within 60-90 min and persisted usually for 8-16 hr. At defervescence, core temperature fell to a level near that of the pre-lesioning base line.3. The prostaglandin synthesis inhibitor, indomethacin, administered I.P. at doses of 5 and 15 mg/kg 1 hr before puncture of the AH/PO region, attenuated the lesion-induced hyperthermia in a dose dependent fashion. The higher dose reduced peak magnitude by 80% and the 6 hr Fever Index by 88%. The vehicle used to dissolve the indomethacin (60% DMSO/40% saline) did not significantly attenuate the hyperthermia.4. In rats that were hyperthermic after AH/PO damage, indomethacin (10-15 mg/kg I.P.) caused core temperature to fall promptly to near the prelesion base line. Reversal occurred whether the indomethacin was injected while core temperature was still rising or late in the plateau phase of the hyperthermia.5. It is suggested that the neurogenic hyperthermia elicited by unilateral lesioning of the AH/PO region was mediated by prostaglandins released from injured tissue and possibly from extravasated blood. Evidence is cited indicating that the most likely sites of action of the released prostaglandins are the surviving portion of the AH/PO region on the punctured side and the intact contralateral AH/PO region.     

Spinal distribution and metabolism of 2'-O-(2-methoxyethyl)-modified oligonucleotides after intrathecal administration in rats

Intrathecal (IT) delivery of antisense oligodeoxynucleotides (ASO) has been used to study the function of specific gene products in spinal nociception. However, a lack of systematic studies on the spinal distribution and kinetics of IT ASO is a major hurdle to the utilization of this technique. In the present study, we injected rats IT with 2'-O-(2-methoxyethyl) modified phosphorothioate ASO (2'-O-MOE ASO) and examined anatomical and cellular location of the ASO in the spinal cord and dorsal root ganglia (DRG) by immunocytochemistry. At 0.5 h after a single IT injection, immunostaining for ISIS 13920 (a 2'-O-MOE ASO targeting h-ras) localized superficially in the lumbar spinal cord, while at 24 h the immunostaining was distributed throughout the spinal cord and was predominantly intracellular. Double staining with cell type specific antibodies indicated that the ASO was taken up by both glia and neurons. ASO immunoreactivity was also observed in DRG after IT ISIS 13920. Capillary gel electrophoresis analysis showed that ISIS 22703, a 2'-O-MOE ASO targeting the alpha isozyme of protein kinase C (PKC), remained intact in spinal cord tissue and cerebrospinal fluid up to 24 h after the injection and no metabolites were detected. In contrast, after IT ISIS 11300, an unmodified phosphorothioate ASO with the same sequence as ISIS 22703, no full-length compound was detectable at 24 h, and metabolites were seen as early as 0.5 h. IT treatment with ISIS 22703 at doses that effectively down-regulated PKCalpha mRNA in spinal cord did not affect the mRNA expression in DRG. In summary, 2'-O-MOE ASO displayed high stability in spinal tissue after IT delivery, efficiently distributed to spinal cord, and internalized into both neuronal and non-neuronal cells. ASO are able to reach DRG after IT delivery; however, higher doses may be required to reduce target gene in DRG as compared with spinal cord.     

Cytochemical, functional, and proliferative characteristics of promonocytes and monocytes from patients with monocytic leukemia

This article deals with a prospective study on the cytochemical, functional, and proliferative characteristics of promonocytes and bone marrow and peripheral blood monocytes of 20 patients with acute monocytic leukemia and 7 patients with chronic monocytic leukemia. The results show a wide variation in the peroxidase and esterase activities in these cells, whereas the percentages of mononuclear phagocytes with Fc gamma and C3b receptors did not differ appreciably from those in normal individuals. A discriminant analysis of these data and corresponding data from normal individuals showed that a below-normal peroxidase activity of circulating monocytes has predictive value for the presence of monocytic leukemia; a below-normal esterase activity has less, but nevertheless some, predictive value in this respect. An increase in the percentage of circulating monocytes, a decrease in the percentage of Fc gamma or C3b receptors, and a decline in the ability to phagocytose bacteria has no predictive value for the presence of monocytic leukemia. The mean percentage of patients' promonocytes that incorporated 3H-thymidine amounted to 80.9%, which is close to the control value in normal individuals. The mean values for the labeling indices of cultured bone marrow and peripheral blood monocytes are 1.0% and 0.74%, respectively; when 3H-thymidine was added to whole blood, the labeling index of the monocytes amounted to 3.6%. These percentages are only a little higher than those found for monocytes of normal individuals. These results indicate that the majority of the circulating monocytes in acute and chronic monocytic leukemia are not actively dividing or blast cells.     

Intrathecal minocycline attenuates peripheral inflammation-induced hyperalgesia by inhibiting p38 MAPK in spinal microglia

Activation of p38 mitogen-activated protein kinase (p38) in spinal microglia is implicated in spinal nociceptive processing. Minocycline, a tetracycline derivative, displays selective inhibition of microglial activation, a function that is distinct from its antibiotic activity. In the present study we examined antinociceptive effects of intrathecal (IT) administration of minocycline in experimental models of inflammation-evoked hyperalgesia in addition to the effect of minocycline on stimulation-induced activation of p38 in spinal microglia. Intrathecal minocycline produced a dose-dependent reduction of formalin-evoked second-phase flinching behaviour in rats, and prevented thermal hyperalgesia induced by carrageenan injection into the paw. In contrast, systemic delivery (intraperitoneally) of minocycline inhibited the first but not the second phase of formalin-induced flinching, and it had no effect on carrageenan-induced hyperalgesia. Centrally mediated hyperalgesia induced by IT delivery of N-methyl-d-aspartate was completely blocked by IT minocycline. An increase in phosphorylation (activation) of p38 (P-p38) was observed in the dorsal spinal cord after carrageenan paw injection, assessed by both Western blotting and immunohistochemistry. The increased P-p38 immunoreactivity was seen primarily in microglia but also in a small population of neurons. Minocycline, at the IT dose that blocked carrageenan-induced hyperalgesia, also attenuated the increased P-p38 in microglia. In addition, minocycline suppressed lipopolysaccharide-evoked P-p38 in cultured spinal microglial cells. Taken together, these findings show that minocycline given IT produces a potent and consistent antinociception in models of tissue injury and inflammation-evoked pain, and they provide strong support for the idea that this effect is mediated by direct inhibition of spinal microglia and subsequent activation of p38 in these cells.     

Increased hyperalgesia after tissue injury and faster recovery of allodynia after nerve injury in the GalR1 knockout mice

Evidence suggests that galanin and its receptors including GalR1 are involved in the modulation of nociception. To understand the contributions of this galanin receptor subtype to the analgesic effect of galanin, we systematically examined the nociception phenotype of the GalR1 knockout (KO) mice. (1) Baseline thresholds: Thermal escape latencies and tactile thresholds of the hind paws were not different between the GalR1 KO and wild type (WT) mice. (2) Thermal injury evoked hyperalgesia: Thermal injury (52 degrees C, 45 s) to one hind paw resulted in a reduction in the thermal escape latency as compared to the uninjured paw. The right/left difference score was significantly greater in the KO (5.9 +/- 0.8 s) than for the WT (2.8 +/- 0.7 s) indicating a greater hyperalgesia. (3) Formalin-induced flinching: Formalin paw injection (2.5%/20 microl) produced a two-phase flinching in both GalR1 KO and WT groups, that was detected by an automated flinching sensor device. Phase II flinching of KO (1510 +/- 90) was slightly greater than that observed for WT (1290 +/- 126), but the difference is not statistically significant. (4) Nerve injury evoked allodynia: Tactile thresholds were assessed prior to and at intervals up to 21 days after left L5 spinal nerve ligation and transection. In both GalR1 KO and WT mice, nerve injury caused thresholds to fall to 0.2-0.3g though 11 days. On days 14-21, GalR1 KO animals showed a significant recovery as compared to WT. In summary, GalR1 KO mice showed no difference from WT with respect to acute nociception, but showed a modest tendency towards increased hyperalgesia after tissue injury and inflammation. These results are consistent with a regulatory effect of galanin at GalR1 receptors on nociceptive processing.     

Prostaglandin E2 release evoked by intrathecal dynorphin is dependent on spinal p38 mitogen activated protein kinase

Spinal dynorphin has been hypothesized to play a pivotal role in spinal sensitization. Although the mechanism of this action is not clear, several lines of evidence suggest that spinal dynorphin-induced hyperalgesia is mediated through an increase in spinal cyclooxygenase products via an enhanced N-methyl-D-aspartate (NMDA) receptor function. Spinal NMDA-evoked prostaglandin release and nociception has been linked to the activation of p38 mitogen activated protein kinase (p38). In the present work, we show that intrathecal delivery of an N-truncated fragment of dynorphin A, dynorphin A 2-17 (dyn2-17), which has no activity at opioid receptors, induced a 8-10-fold increase in phosphorylation of p38 in the spinal cord. The increase in phosphorylated p38 was detected in laminae I-IV of the dorsal horn. Moreover, confocal microscopy showed that the activation of p38 occurred in microglia, but not in neurons or astrocytes. In awake rats, prepared with chronically placed intrathecal loop dialysis catheters, the concentration of prostaglandin E2 in lumbar cerebrospinal fluid was increased 5-fold by intrathecal administration of dyn2-17. Injection of SD-282, a selective p38 inhibitor, but not PD98059, an ERK1/2 inhibitor, attenuated the prostaglanin E2 release. These data, taken together, support the hypothesis that dynorphin, independent of effects mediated by opioid receptors, has properties that can induce spinal sensitization and indicates that dyn2-17 effects may be mediated through activation of the p38 pathway. These studies provide an important downstream linkage where by dynorphin may act through a non-neuronal link to induce a facilitation of spinal nociceptive processing.