Release Of CCK-LI Research Articles

Arthritis-induced increase in cholecystokinin release in the rat anterior cingulate cortex is reversed by diclofenac

Given a hypothesised role for CCK in the anterior cingulate cortex (ACC) for the sensation of pain, the aim of the present study was to investigate whether the increased CCK release could be affected by two different analgesic drugs, morphine and the non-selective cyclooxygenase inhibitor diclofenac. Since opioids stimulate CCK release in other CNS regions we have also studied the effect of morphine by itself on the CCK-LI release in the ACC of non-arthritic rats. Three to seven hours after intraarticular carrageenan injection, at the time when the animals are known to show pain-related behaviour, in vivo microdialysis in awake rats revealed increased CCK-LI release in the ACC. The CCK-LI release was significantly attenuated by diclofenac (25 mg/kg i.m.), but not by morphine (10 mg/kg s.c.). Neither diclofenac (25 mg/kg i.m.) nor morphine (5 or 10 mg/kg s.c.) affected the CCK-LI release in the ACC in non-arthritic rats. The results obtained with diclofenac indicate that prostaglandins contribute to the increased CCK-LI release in the ACC during monoarthritis. However, the lack of effect of morphine suggests that the CCK release in the ACC is not directly related to the sensation of pain. Further on, the failure of morphine to affect the extracellular level of CCK-LI in the ACC in control animals as well as in animals with carrageenan-induced monoarthritis is in contrast to previous studies on the frontal cortex or the dorsal horn of the spinal cord, in which similar doses of morphine stimulate CCK release. Thus, compared to these regions, CCK release may be differently regulated in the ACC.

Increased cholecystokinin release in the rat anterior cingulate cortex during carrageenan-induced arthritis

Several human and animal studies indicate that the anterior cingulate cortex (ACC) plays an important role in the affective component of pain. The neuropeptide cholecystokinin (CCK) is especially abundant in the ACC. CCK has been suggested to be involved in the mediation of anxiety and in the modulation of opioid effects in the spinal cord and medulla oblongata. However, its possible role in pain transmission or modulation in the brain is far less clear. In this study, a model of subchronic inflammatory pain in rats, carrageenan-induced monoarthritis, was used to study the effect of pain on the release of CCK-like immunoreactivity (CCK-LI) in the ACC. Pain-related behaviour quantified by weight bearing and stance scoring, as well as inflammation measured by ankle oedema, was increased for at least 24 h after carrageenan injection with a maximum at 5 h. Using microdialysis in freely moving rats, extracellular concentrations of CCK-LI was measured in the ACC during a time period when the animals showed significant pain behaviour. In animals with carrageenan-induced arthritis, both basal and potassium-evoked release of CCK-LI were significantly increased compared to controls. HPLC analysis of dialysates from the ACC during potassium stimulation showed that the main part of the immunoreactive material was sulphated CCK-8. Because CCK has been implicated in anxiety, we suggest that an altered CCK-ergic activity in the ACC may be of importance for the affective component of pain, but an involvement in the modulation of nociception is also possible.

External pH changes affect NMDA-evoked and spontaneous release of cholecystokinin, somatostatin and noradrenaline from rat cerebrocortical nerve endings

It was previously reported that the K +-evoked release of somatostatin-like immunoreactivity (SRIF-LI) and of cholecystokinin-like immunoreactivity (CCK-LI) from superfused rat cerebrocortical synaptosomes can be enhanced by NMDA or d-serine alone. We here studied the effects of extraterminal pH changes on SRIF-LI and CCK-LI release. Lowering pH from 7.4 to 6.9 or 6.4 abolished the effects of NMDA or d-serine on the K +-evoked peptide release. Identical results were obtained when external pH was raised to 8 or 8.7. Sudden alkalinization of the superfusion medium, in absence of K +-depolarization, induced SRIF-LI or CCK-LI release which was insensitive to NMDA. Based on experiments in Ca 2+-free medium and with voltage-sensitive Ca 2+ channel (VSCC) blockers, the pH 8.7-induced release of SRIF-LI and CCK-LI was only in part (30–50%) dependent on external Ca 2+ and Ca 2+ channel activation. In contrast, the alkalinization-evoked release of [ 3 H ]noradrenaline was highly sensitive to external Ca 2+ removal and to blockade of Ca 2+ channels with ω-conotoxins. The pH 8.7-evoked SRIF-LI and CCK-LI was about halved in synaptosomes intoxicated with botulinum toxin C1. The results suggest that the pH-sensitive NMDA receptors mediating somatostatin and cholecystokinin release contain NR1 subunits lacking the exon-5 cassette. Alkalinization represents a novel releasing stimulus which elicits neuropeptide release in part by conventional exocytosis and largely by an external Ca 2+-independent mechanism. Differently, the release of noradrenaline provoked by alkalinization occurs entirely by conventional exocytosis.

External pH changes affect NMDA-evoked and spontaneous release of cholecystokinin, somatostatin and noradrenaline from rat cerebrocortical nerve endings

It was previously reported that the K + -evoked release of somatostatin-like immunoreactivity (SRIF-LI) and of cholecystokinin-like immunoreactivity (CCK-LI) from superfused rat cerebrocortical synaptosomes can be enhanced by NMDA or d -serine alone. We here studied the effects of extraterminal pH changes on SRIF-LI and CCK-LI release. Lowering pH from 7.4 to 6.9 or 6.4 abolished the effects of NMDA or d -serine on the K + -evoked peptide release. Identical results were obtained when external pH was raised to 8 or 8.7. Sudden alkalinization of the superfusion medium, in absence of K + -depolarization, induced SRIF-LI or CCK-LI release which was insensitive to NMDA. Based on experiments in Ca 2+ -free medium and with voltage-sensitive Ca 2+ channel (VSCC) blockers, the pH 8.7-induced release of SRIF-LI and CCK-LI was only in part (30–50%) dependent on external Ca 2+ and Ca 2+ channel activation. In contrast, the alkalinization-evoked release of [ 3 H ]noradrenaline was highly sensitive to external Ca 2+ removal and to blockade of Ca 2+ channels with ω-conotoxins. The pH 8.7-evoked SRIF-LI and CCK-LI was about halved in synaptosomes intoxicated with botulinum toxin C1. The results suggest that the pH-sensitive NMDA receptors mediating somatostatin and cholecystokinin release contain NR1 subunits lacking the exon-5 cassette. Alkalinization represents a novel releasing stimulus which elicits neuropeptide release in part by conventional exocytosis and largely by an external Ca 2+ -independent mechanism. Differently, the release of noradrenaline provoked by alkalinization occurs entirely by conventional exocytosis.

Pharmacological characterization of morphine-induced in vivo release of cholecystokinin in rat dorsal horn: effects of ion channel blockers.

Previous studies indicate that an increased release of cholecystokinin (CCK) in response to morphine administration may counteract opioid-induced analgesia at the spinal level. In the present study we used in vivo microdialysis to demonstrate that systemic administration of antinociceptive doses of morphine (1-5 mg/kg, s.c.) induces a dose-dependent and naloxone-reversible release of CCK-like immunoreactivity (CCK-LI) in the dorsal horn of the spinal cord. A similar response could also be observed following perfusion of the dialysis probe for 60 min with 100 microM but not with 1 microM morphine. The CCK-LI release induced by morphine (5 mg/kg, s.c.) was found to be calcium-dependent and tetrodotoxin-sensitive (1 microM in the perfusion medium). Topical application of either the L-type calcium channel blocker verapamil (50 microg) or the N-type calcium channel blocker omega-conotoxin GVIA (0.4 microg) onto the dorsal spinal cord completely prevented the CCK-LI release induced by morphine (5 mg/kg, s.c.). Our data indicate that activation of L- and N-type calcium channels is of importance for morphine-induced CCK release, even though the precise site of action of morphine in the dorsal horn remains unclear. The present findings also suggest a mechanism for the potentiation of opioid analgesia by L- and N-type calcium channel blocking agents.

Morphine-induced in vivo release of spinal cholecystokinin is mediated by delta-opioid receptors--effect of peripheral axotomy.

Morphine and other opioid agonists induce spinal in vivo release of cholecystokinin (CCK), a neuropeptide with anti-opioid properties. However, so far the opioid receptor subtype responsible for this effect has not been determined. In the present in vivo microdialysis study, the morphine-induced release of cholecystokinin-like immunoreactivity (CCK-LI) in the dorsal horn was completely blocked by the delta-opioid antagonist naltrindole (10 microM in the perfusion fluid). Neither the mu-opioid receptor antagonist D-Phe-Cys-Tyr-D-Trp-Orn-Thr-Pen-Thr amide (CTOP; 10 microM in the perfusion fluid), nor the kappa-opioid receptor antagonist nor-binaltorphimine (nor-BNI); 10 microM in the perfusion fluid) had any significant effect in this respect. In addition, systemic administration of the delta-opioid receptor agonist BW373U86 (1 mg/kg, s.c.) and spinal administration of the delta(2)-opioid receptor agonist, Tyr-D-Ala-Phe-Glu-Val-Val-Gly amide ([D-Ala(2)] deltorphin II) (1 microM in the perfusion fluid) induced a significant increase of the CCK-LI level. The effect of BW373U86 on spinal CCK-LI release was completely blocked by spinal administration of naltrindole. The mu-opioid receptor agonist [D-ala(2)-N-Me-Phe(4)-Gly(5)-ol]-enkephalin (DAMGO) (1 microM in the perfusion fluid or 1 mg/kg, s.c.) failed to alter the CCK-LI level. Peripheral nerve lesions have previously been shown to down-regulate mu- and delta-opioid receptors in the dorsal horn, to increase the gene-expression of CCK and CCK-receptor mRNA in dorsal root ganglion neurons and to alter the potassium-induced spinal CCK-LI release. After complete sciatic nerve transection, administration of the two selective delta-opioid receptor agonists induced a significant release of CCK-LI, which was comparable to controls. In contrast, neither systemic nor spinal administration of morphine and DAMGO altered the spinal CCK-LI release in axotomized animals. The present data indicate that the delta-opioid receptor mediates morphine-induced CCK-LI release in the spinal cord.

Changes in spinal cholecystokinin release after peripheral axotomy.

The gene expression of cholecystokinin (CCK), a neuropeptide with anti-opioid properties, has been reported to be upregulated in some primary sensory neurons after a peripheral nerve lesion. We have recently demonstrated that the upregulation of CCK mRNA is not accompanied by an increased potassium-evoked release CCK-like immunoreactivity (CCK-LI) 2-4 weeks after a complete transection of the sciatic nerve. The potassium-evoked release of CCK-LI at earlier and later time points has, however, not been studied. The aim of the present in vivo microdialysis study was to monitor how the basal and stimulated extracellular level of CCK in the dorsal horn of the spinal cord is affected at various time points after a complete transection of the sciatic nerve (axotomy). During the first week after transection of the sciatic nerve a tendency towards an elevation of the potassium-induced (100 mM in the perfusion fluid) release of spinal CCK-LI was observed. In contrast, no potassium-induced release of CCK-LI could be detected 2-3 weeks and 2 months after axotomy. No significant effect was observed on the basal extracellular levels of CCK-LI in the dorsal horn. The present study provides further support for the notion that the adaptive changes in the dorsal horn 2 weeks and later after a deafferentiation injury do not include an increased release of CCK.

The HIV-1 coat protein gp120 and some of its fragments potently activate native cerebral NMDA receptors mediating neuropeptide release.

The objective of this study was to investigate the effects of the HIV-1 envelope protein gp120 and its peptide fragments on the function of N-methyl-D-aspartate (NMDA) receptors mediating release of cholecystokinin (CCK) and somatostatin (SRIF). These are nonconventional NMDA receptors recently found to be activated by glycine or D-serine 'only'. The release of cholecystokinin-like immunoreactivity (CCK-LI) and of somatostatin-like immunoreactivity (SRIF-LI) elicited by 12 mM K+ from superfused rat neocortex synaptosomes was potently increased by gp120, its cyclic V3 loop and the linear V3 sequence BRU-C-34-A, but not by RP-135 (a central portion of BRU-C-34-A). The EC50 values of gp120 were 0.02 nM (CCK-LI release) and 0.01 nM (SRIF-LI release). The releasing effect of gp120 was prevented by blocking the glycine site or the ion channel of NMDA receptors, but not the glutamate recognition site; in addition, the gp120 effect was strongly inhibited by nanomolar concentrations of Zn2+ ions and by low micromolar concentrations of ifenprodil. It is concluded that gp120 acts as a very potent agonist at the glycine site of NMDA receptors sited on CCK- and SRIF-releasing nerve endings; the protein is able to activate the receptor channel in the absence of glutamate. Gp120 activates the receptors through its V3 loop as peptide fragments related to V3 retain near-maximal activity. The sensitivity of the gp120 effect to both Zn2+ and ifenprodil would not be incompatible with the idea that these NMDA receptors contain the triple subunit combination NR1/NR2A/NR2B.

Evidence for functional native NMDA receptors activated by glycine or D-serine alone in the absence of glutamatergic coagonist.

In this study we have examined the effects of N-methyl-D-aspartate (NMDA) receptor activation on the release of cholecystokinin and somatostatin from rat neocortical nerve endings. The release of cholecystokinin-like immunoreactivity (CCK-LI) and of somatostatin-like immunoreactivity (SRIF-LI) elicited by 12 mM K+ from superfused synaptosomes, but not the spontaneous release, was increased by NMDA in a concentration-dependent manner. The effects of NMDA could be prevented by antagonists selective for the glutamate recognition site, the receptor channel and the glycine site of the NMDA receptor. In the absence of NMDA, glycine increased on its own and in a concentration-dependent manner the depolarization-evoked release of both CCK-LI and SRIF-LI. This effect of glycine was strychnine-insensitive and could be mimicked by D-serine, a stereoselective agonist at the NMDA receptor glycine site. Antagonists selective for the glycine site or for the NMDA receptor channel prevented the effects of glycine/D-serine; these effects were, however, insensitive to blockade of the glutamate recognition site of the NMDA receptor, suggesting that glutamate released from synaptosomes or present as contaminant was not involved. The neuropeptide release elicited by D-serine was strongly inhibited by ifenprodil (0.3 microM) and by Zn2+ ions (50 nM), selective ligands at the NR2B and NR2A subunits of NMDA receptors, respectively. It is concluded that nerve terminals of CCK- and SRIF-releasing neurons possess non-conventional NMDA receptors whose channels can be operated by glycine or D-serine without apparent activation of the glutamatergic coagonist site. These receptors may display the triple subunit combination NR1/NR2A/NR2B.

Peripheral axotomy influences the in vivo release of cholecystokinin in the spinal cord dorsal horn—possible involvement of cholecystokinin-B receptors

An increased expression of cholecystokinin (CCK) messenger RNA (mRNA) as well as CCK-B receptor mRNA in dorsal root ganglion (DRG) cells following peripheral axotomy has previously been demonstrated. In the present in vivo microdialysis study, the effect of unilateral sciatic nerve section on basal and potassium-induced release of CCK-like (CCK-LI) immunoreactivity in the rat dorsal horn was investigated. We also compared the effects of the CCK-B receptor antagonist CI988 on basal and potassium-stimulated CCK-LI release in intact animals and in chronically axotomized rats. Perfusion of the microdialysis probe with KCl (100 mM) induced a more than 6-fold increase of the extracellular level of CCK-LI in control animals. In contrast, following unilateral sciatic nerve section the same KCl stimulation failed to evoke a release of CCK-LI ipsilaterally. However, after systemic administration of CI988 (1 mg kg −1, i.v.), 100 mM KCl induced a significant increase of the extracellular CCK-LI level in axotomized rats, similar to that observed in control animals. In control animals no effect of CI988 on KCl-stimulated CCK-LI release could be detected. CI988 by itself had no influence on the extracellular CCK-LI level in either nerve injured or control animals. The present data suggest that axotomy reduces the release of CCK-like immunoreactivity in the spinal cord by a mechanism involving the CCK-B receptor binding site.

508 Studies on the centrally administered nitric oxide synthesis inhibitor on the cardiovascular regulation system

An increased expression of cholecystokinin (CCK) messenger RNA (mRNA) as well as CCK-B receptor mRNA in dorsal root ganglion (DRG) cells following peripheral axotomy has previously been demonstrated. In the present in vivo microdialysis study, the effect of unilateral sciatic nerve section on basal and potassium-induced release of CCK-like (CCK-LI) immunoreactivity in the rat dorsal horn was investigated. We also compared the effects of the CCK-B receptor antagonist CI988 on basal and potassium-stimulated CCK-LI release in intact animals and in chronically axotomized rats. Perfusion of the microdialysis probe with KCl (100 mM) induced a more than 6-fold increase of the extracellular level of CCK-LI in control animals. In contrast, following unilateral sciatic nerve section the same KCl stimulation failed to evoke a release of CCK-LI ipsilaterally. However, after systemic administration of CI988 (1 mg kg−1, i.v.), 100 mM KCl induced a significant increase of the extracellular CCK-LI level in axotomized rats, similar to that observed in control animals. In control animals no effect of CI988 on KCl-stimulated CCK-LI release could be detected. CI988 by itself had no influence on the extracellular CCK-LI level in either nerve injured or control animals. The present data suggest that axotomy reduces the release of CCK-like immunoreactivity in the spinal cord by a mechanism involving the CCK-B receptor binding site.

Studies on the centrally administered nitric oxide synthesis inhibitor on the cardiovascular regulation system

An increased expression of cholecystokinin (CCK) messenger RNA (mRNA) as well as CCK-B receptor mRNA in dorsal root ganglion (DRG) cells following peripheral axotomy has previously been demonstrated. In the present in vivo microdialysis study, the effect of unilateral sciatic nerve section on basal and potassium-induced release of CCK-like (CCK-LI) immunoreactivity in the rat dorsal horn was investigated. We also compared the effects of the CCK-B receptor antagonist CI988 on basal and potassium-stimulated CCK-LI release in intact animals and in chronically axotomized rats. Perfusion of the microdialysis probe with KCl (100 mM) induced a more than 6-fold increase of the extracellular level of CCK-LI in control animals. In contrast, following unilateral sciatic nerve section the same KCl stimulation failed to evoke a release of CCK-LI ipsilaterally. However, after systemic administration of CI988 (1 mg kg−1, i.v.), 100 mM KCl induced a significant increase of the extracellular CCK-LI level in axotomized rats, similar to that observed in control animals. In control animals no effect of CI988 on KCl-stimulated CCK-LI release could be detected. CI988 by itself had no influence on the extracellular CCK-LI level in either nerve injured or control animals. The present data suggest that axotomy reduces the release of CCK-like immunoreactivity in the spinal cord by a mechanism involving the CCK-B receptor binding site.

Effect of chemical kindling model induced by PTZ on CCK mRNA and CCK-LI levels in rat brain regions

An increased expression of cholecystokinin (CCK) messenger RNA (mRNA) as well as CCK-B receptor mRNA in dorsal root ganglion (DRG) cells following peripheral axotomy has previously been demonstrated. In the present in vivo microdialysis study, the effect of unilateral sciatic nerve section on basal and potassium-induced release of CCK-like (CCK-LI) immunoreactivity in the rat dorsal horn was investigated. We also compared the effects of the CCK-B receptor antagonist CI988 on basal and potassium-stimulated CCK-LI release in intact animals and in chronically axotomized rats. Perfusion of the microdialysis probe with KCl (100 mM) induced a more than 6-fold increase of the extracellular level of CCK-LI in control animals. In contrast, following unilateral sciatic nerve section the same KCl stimulation failed to evoke a release of CCK-LI ipsilaterally. However, after systemic administration of CI988 (1 mg kg−1, i.v.), 100 mM KCl induced a significant increase of the extracellular CCK-LI level in axotomized rats, similar to that observed in control animals. In control animals no effect of CI988 on KCl-stimulated CCK-LI release could be detected. CI988 by itself had no influence on the extracellular CCK-LI level in either nerve injured or control animals. The present data suggest that axotomy reduces the release of CCK-like immunoreactivity in the spinal cord by a mechanism involving the CCK-B receptor binding site.

Simultaneous determination of cholecystokinin-like immunoreactivity and dopamine release after treatment with veratrine, NMDA, scopolamine and SCH23390 in rat medial frontal cortex: a brain microdialysis study.

The release of cholecystokinin (CCK) and dopamine (DA) was measured simultaneously in the medial frontal cortex of the rat using an in vivo microdialysis technique coupled to either enzyme immunoassay or high performance liquid chromatography with electrochemical detection method. Basal levels of CCK-like immunoreactivity (CCK-LI) and DA in the dialysates were 0.29 +/- 0.05 pg/50 microliters and 4.98 +/- 0.56 fmol/20 microliters respectively. After perfusion of 100 micrograms/ml veratrine into rat medial frontal cortex through the microdialysis probe, the release of those neurotransmitters was significantly enhanced. Perfusion of 10 mM NMDA induced significant increases of CCK-LI and DA levels. Co-perfusion of 1 mM SCH23390, a D1 receptor antagonist, suppressed NMDA-evoked CCK-LI release and slightly raised NMDA-evoked DA release. Treatment with scopolamine, a muscarinic receptor antagonist, suppressed veratrine-induced CCK-LI and DA release, but did not change NMDA-evoked CCK-LI and DA release. These results suggest that NMDA may regulate CCK-LI release through the activation of D1 receptor existing on the afferent CCK neurons or interneurons in rat medial frontal cortex, and changes of endogenous CCK release may be involved in NMDA receptor-mediated long-term potentiation.

Inhibition by 5-HT3 receptor antagonists of release of cholecystokinin-like immunoreactivity from the frontal cortex of freely moving rats.

Effects of the 5-HT3 receptor antagonists, ondansetron and tropisetron, on the release of cholecystokinin-like immunoreactivity (CCK-LI) in rat frontal cortex were investigated in conscious, unrestrained rats using intracerebral microdialysis. The release of CCK-LI was augmented by perfusion with 100 micrograms/ml veratrine and was fully Ca(2+)-dependent and tetrodotoxin-sensitive. Ondansetron and tropisetron, each at 0.1-1 mumol/l, decreased concentration-dependently the veratrine-evoked efflux of CCK-LI. The reduction of CCK-LI output was approximately 30% when the antagonists were infused at 0.1 mumol/l. The data suggest that 5-HT3 receptor antagonists prevent the release of CCK evoked by endogenous 5-hydroxytryptamine. These drugs may thus represent a novel therapeutic approach in disease states, like anxiety, in which an inappropriately high release of brain CCK or 5-hydroxytryptamine seems to be involved.

Role of cyclic nucleotides and calcium in the nutrient-induced release of cholecystokinin-like immunoreactivity in rats.

1. This study was undertaken with an isolated vascularly perfused rat duodenojejunal preparation to investigate the mechanisms of the release of cholecystokinin measured by immunoassay (cholecystokinin-like immunoreactivity, CCK-LI). 2. Intra-arterial infusion of forskolin (2-20 microM) evoked a prompt and well-sustained secretion of CCK-LI which was increased to a mean value of 600% of basal with the highest dose tested. 3-Isobutyl-1-methylxanthine (IBMX) (10(-6)-10(-4) M) stimulated the secretion of CCK-LI (maximal increase of 400% of basal at 10(-4) M). 3. Intra-arterial infusion of beta-phorbol 12-myristate 13-acetate (5 x 10(-7)-5 x 10(-6) M) and calcium ionophore A23187 (10(-6)-10(-5) M) alone or in combination provoked only a transient increase in the release of CCK-LI. 4. Luminal infusion of a 5% ovalbumin hydrolysate solution produced an immediate release of CCK-LI followed by a well-sustained secretion at 580% of basal. Intra-arterial infusion of IBMX (10(-5) or 10(-4) M) in combination with luminal peptone induced a release of CCK-LI which was equal to the sum of the CCK responses evoked by IBMX and peptone given separately. 5. Intra-arterial infusion of EGTA (2 mM) abolished the forskolin- and peptone-induced CCK secretion while luminal EGTA (2 mM) had no inhibitory effect. Verapamil (5 x 10(-5)-10(-4) M) or nifedipine (10(-5)-5 x 10(-5) M) inhibited the peptone-evoked CCK secretion. A high concentration of trifluoperazine (10(-4) M) strongly reduced the release of CCK-LI induced by intraluminal peptone while 10(-5) M was ineffective. 6. It is concluded that the peptone-induced secretion of CCK-LI involves a cyclic AMP-dependent mechanism and the activation of calcium channels possibly located at the basolateral side of the CCK cell.

Cholecystokinin release mediated by 5‐HT3 receptors in rat cerebral cortex and nucleus accumbens

1. The effects of 5-hydroxytryptamine (5-HT) on the release of cholexystokinin-like immunoreactivity (CCK-LI) were examined in synaptosomes prepared from rat cerebral cortex and nucleus accumbens and depolarized by superfusion with 15 mM KCl. 2. In both areas 5-HT, tested between 0.1 and 100 nM, increased the calcium-dependent, depolarization-evoked CCK-LI release in a concentration-related manner. The concentration-response curves did not differ significantly between the two brain areas (EC50: 0.4 +/- 0.045 nM and 0.48 +/- 0.053 nM, respectively, in cortical and n. accumbens synaptosomes; maximal effect: about 60% at 10 nM 5-HT). 3. The 5-HT1/5-HT2 receptor antagonist methiothepin (300 nM) did not affect the CCK-LI release elicited by 10 nM 5-HT. However, the effects of 10 nM 5-HT were antagonized in a concentration-dependent manner by the 5-HT3 receptor antagonists (3 alpha-tropanyl)-1H-indole-3-carboxylic acid ester (ICS 205-930; 0.1-100 nM; IC50: 3.56 +/- 0.42 nM in the cortex and 3.90 +/- 0.50 nM in the n. accumbens) and ondasetron (IC50: 8.15 +/- 0.73 nM in the cerebral cortex). 5-HT (10 nM) was also strongly antagonized by 100 nM 1 alpha H, 3 alpha 5 alpha H-tropan-3-yl-3,5-dichlorobenzoate (MDL 72222) another blocker of the 5-HT3 receptor. Moreover, the 5-HT3 receptor agonist 1-phenylbiguanide (tested in the cerebral cortex between 0.1 and 100 nM) enhanced CCK-LI release in a manner almost identical to that of 5-HT (EC50 = 0.64 +/- 0.071 nM). 4. It is concluded that 5-HT can act as a potent releaser of CCK-LI in rat cerebrocortex and nucleus accumbens through the activation of receptors of the 5-HT3 type situated on the CCK-releasing terminals. This interaction may provide a rationale for the clinical development of both 5-HT3 and CCK receptor antagonists as novel anxiolytic drugs.

Effect of low-frequency electric stimulation on in vivo release of cholecystokinin-like immunoreactivity in medial thalamus of conscious rat

Release of cholecystokinin-like immunoreactivity (CCK-LI) in the medial thalamus of conscious rats was measured by brain dialysis and enzyme immunoassay. Analgesia caused by low-frequency electric stimulation of the tibial muscle, the tsusanli acupuncture point, was judged by change of pain threshold due to the stimulation. Medial thalamic CCK-LI release was increased by peripheral electric stimulations of both the acupuncture point and the non-acupuncture point. Results suggest that CCK acts as a neurotransmitter in the medial thalamus, a part of the analgesia inhibitory system.

Luminal CCK-releasing factors in the isolated vascularly perfused rat duodenojejunum

The factors operating at the apical side of the endocrine cell releasing cholecystokinin (CCK) were investigated using the isolated vascularly perfused rat duodenojejunum. In the protease-free intestinal segment, a 30-min infusion of glucose (280 mM), oleic acid (100 mM), or triglycerides containing short- or long-chain fatty acids did not alter significantly the basal level of portal CCK-like immunoreactivity (CCK-LI), while octanoic acid (100 mM) produced a transient rise of plasma CCK-LI to approximately 250% of basal. Infusion of proteins (5% solutions of ovalbumin or casein) or of a mixture of all amino acids brought about a modest CCK secretion. In contrast, isocaloric amounts of an ovalbumin hydrolysate produced a sharp rise of portal CCK-LI to 530% of basal followed by a well-sustained plateau secretion (420% of basal) until the end of the infusion. An acid casein hydrolysate induced a slightly less pronounced CCK-LI release and was followed in decreasing order by meat, casein, and soybean peptones. Simultaneous infusion of trypsin with ovalbumin or casein hydrolysate reduced by approximately 60% the CCK release induced by peptone alone. This effect was reversed by coinfusion of soybean trypsin inhibitor (SBTI) with the trypsin-peptone mixture. Arterial infusion of tetrodotoxin (10(-6) M) or atropine (10(-5) M) had no significant effect on the trypsin-induced inhibition of peptone-mediated CCK-LI release. Administration of SBTI or camostate alone or in combination with trypsin did not alter basal CCK. Monitor peptide produced a dose-dependent transient rise of portal CCK-LI over the range from 2 to 12 micrograms.(ABSTRACT TRUNCATED AT 250 WORDS)

Blockade of bombesin receptors with [Leu 14-ψ(CH 2NH)-Leu 13]bombesin fails to suppress nutrient-induced CCK release from rat duodenojejunum

The present study was undertaken in order to delineate the contribution of enteric bombesin (BBS)-containing nerves in the food-induced release of intestinal cholecystokinin (CCK). For this purpose, the isolated vascularly perfused rat duodenojejunum model was used and the new compound [Leu 14-ψ(CH 2NH)-Leu 13]BBS was infused intraarterially at a concentration of 10 −6 M to block the BBS receptors. Vascular infusion of BBS alone (10 −8 M or 10 −9 M) provoked a dose-dependent release of CCK-like immunoreactivity (CCK-LI). The secretion pattern of CCK was biphasic and consisted of a transient peak (300–400% above basal) followed by a sustained response (200–300% above basal). Vascular coinfusion of the BBS analogue with BBS 10 −9 M completely abolished both phases of CCK release while only the second phase of CCK secretion was profoundly reduced upon coadministration of BBS 10 −8 M with the BBS receptor antagonist. Luminal administration of mixed nutrients induced a prompt and well-sustained release of CCK-LI which was unaffected upon arterial infusion of the BBS analogue. These data suggest that the intestinal supply in BBS-producing nerves is not involved in the food-induced release of intestinal CCK in the rat.