Inhibition of electroshock-induced seizures by cholecystokinin-related peptides in mice

The effects of several doses of cholecystokinin octapeptide sulphate ester (CCK-8-SE) and nonsulphated cholecystokinin octapeptide (CCK-8-NS), and two CCK-related peptide analogues Ac-Thr5-caerulein, and nonsulphated Ac-Thr5-caerulein were investigated on electroshock-(ES)-induced seizures after intraperitoneal administration in mice. As parameters, the duration of the tonic and clonic phase of the fit, and those of postictal coma and behavioural depression were measured. CCK-8-SE decreased the duration of the clonic phase; its highest dose, 3.2 mumol/kg, shortened the coma. CCK-8-NS antagonized only slightly the clonic phase of seizure. Ac-Thr5-caerulein did not influence ES-induced seizures in any dose, only increased the duration of behavioural depression. Similarly to CCK-8-NS, the nonsulphated form of Ac-Thr5-caerulein inhibited selectively the clonic phase of seizures. The reference drugs, diazepam and phenobarbital, antagonized dose-dependently and most effectively the tonic phase of ES-induced seizures, but in much higher doses than did the CCK-related peptides. Besides, diazepam increased and phenobarbital decreased the duration of postictal coma. The results showed that the tested CCK-related peptides inhibit prevalently the clonic phase of ES-induced seizures after peripheral administration.     

The inhibition of tail-pinch-induced food intake by cholecystokinin octapeptides and their fragments

The effects of intraperitoneally (ip.) and intracerebroventricularly (icv.) administered sulfated and nonsulfated cholecystokinin octapeptide (CCK-8-SE and CCK-8-NS) and their N- and C-terminal fragments on the tail-pinch-induced feeding behavior of rats were investigated. After ip. administration, only CCK-8-SE inhibited tail-pinch-induced food intake. After icv. administration, both CCK-8-SE and CCK-8-NS, in doses of 800 pmole/rat, reduced the amount of food eaten. Of the CCK fragments tested icv., the sulfated N-terminal fragments, the middle portion of the CCK-8-sequence (the CCK-3-6 fragment), and the C-terminal tetrapeptide depressed the food intake of rats during tail-pinch, whereas the C-terminal tripeptide significantly increased it. The results suggest that CCK peptides inhibit tail-pinch-induced feeding by separate mechanisms, depending on the route of administration.     

Effects of cholecystokinin-related peptides on retention of passive avoidance behaviour

Effects of intraperitoneal administration of cholecystokinin (CCK)-related peptides were studied on retention of single-trial learning passive avoidance behaviour. The COOH-terminal octapeptide of CCK (CCK-1-8-SE), the unsulfated octapeptide (CCK-1-8-NS), as well as the COOH-terminal tetrapeptide of CCK (CCK-5-8), administered immediately after the learning trial, facilitated passive avoidance behaviour. The data indicate that these peptides may influence memory consolidation processes.     

Species differences in the behavioral effects of cerulein--an agonist of the receptors of the octapeptide cholecystokinin--in white mice and rats

It has been shown in the behavioural experiments that combined pretreatment with haloperidol (0.25 mg/kg) and caerulein (40 micrograms/kg), and to a lesser extent pretreatment with caerulein alone caused long-term reversal of amphetamine (2 mg/kg) induced hyperexcitability in rats. Administration of proglumide (50 mg/kg), an antagonist of CCK-8 receptors, did not reverse long-term antiamphetamine effect of caerulein. In mice pretreatment with caerulein (50 and 100 micrograms/kg) alone or in combination with haloperidol (0.25 mg/kg) caused hypersensitivity to the behavioural effect of amphetamine (3 mg/kg). Intraventricular (I ng), but not systemic (100-500 micrograms/kg) administration of caerulein selectively antagonized seizures in mice induced by intraventricular administration of quinolinic acid (5 micrograms) and N-methyl-D-aspartate (0.2 microgram). Pretreatment with proglumide (50 mg/kg) reversed the anticonvulsive effect of caerulein in mice. In rats, caerulein failed to affect the seizures caused by intraventricular administration of quinolinic acid. The results of the present study demonstrate the existence of obvious interspecies differences in the behavioural effects of caerulein, the agonist of CCK-8 receptors, in mice and rats.     

Modulation of passive avoidance behaviour of rats by intracerebroventricular administration of cholecystokinin octapeptide sulfate ester and nonsulfated cholecystokinin octapeptide

The effects of intracerebroventricular administration of different doses of cholecystokinin octapeptide sulfate ester (CCK-8-SE) and nonsulfated cholecystokinin octapeptide (CCK-8-NS) were tested on the latency of passive avoidance behaviour in rats. Treatments were carried out prior to learning trial, immediately after electroshock and prior to testing 24 h retention. Both CCK-8-NS and CCK-8-SE enhanced the latency of passive avoidance after all forms of treatment while showing different dose-response patterns depending on time of administration. These data indicate that CCK-8-SE and CCK-8-NS might play a role in the regulation of memory consolidation and retrieval.     

Effects of intraventricular administration of cholecystokinin octapeptide sulfate ester and unsulfated cholecystokinin octapeptide on active avoidance and conditioned feeding behaviour of rats

The effects of cholecystokinin octapeptide sulfate ester (CCK-8-SE) and unsulfated cholecystokinin (CCK-8-NS) were studied following intraventricular administration on active avoidance and conditioned feeding behaviour of rats. In the CCK-8-NS and CCK-8-SE treated animals the acquisition of active avoidance and conditioned feeding behaviour were considerably impaired compared to the control; furthermore, these peptides caused a facilitated extinction of active avoidance and conditioned feeding behaviour. The data suggest that cholecystokinin octapeptide is capable of modifying the fear and hunger motivated behaviours of rats.     

Effects of cholecystokinin octapeptide (CCK-8) on food intake in adult and aged rats under different feeding conditions

The effects of CCK on food intake were investigated under fixed feeding conditions in comparison to a test meal taken after 16 h of food deprivation. The experiments were performed on young adult rats (8 weeks old) as well on aged rats (23 months old). Intraperitoneal CCK-8 (8 and 40 μg/kg) significantly reduced the size of a test meal following 16-h food deprivation. This effect was independent of the age of the rats. However, under fixed feeding conditions neither of the doses used in this study reduced food intake in the young adult rats, whereas the highest dose of 40 μg/kg did so in the aged rats. These results suggest that the hypophagic effect of exogenous CCK-8 depends on experimental conditions, food intake being reduced after a period of food deprivation but not under a fixed feeding regimen in adult animals. Furthermore, the data suggest that age is a factor contributing to the complex behavioral actions of CCK, because only old animals were more susceptible to an anorectic action of CCK under the fixed feeding schedule. An explanation may lie in an interaction of other known behavioral effects of CCK (e.g., anxiogenic, mnemonic action) with its effects under the different feeding schedules.     

Effect of centrally administered cholecystokinin on feeding behavior induced by electric stimulation of the lateral hypothalamus in the rabbit

In experiments on 15 freely moving rabbits cholecystokinin octapeptide sulfate (CCK-8-S) in a dose of 10 ng considerably suppressed alimentary behaviour of the animals elicited by electrical stimulation of the lateral hypothalamus. Increasing of the peptide doses to 100 and 200 ng elicited an analogous effect. CCK-8-NS in 10 ng dose produced a lesser effect on feeding of the animal, but increasing of the dose of nonsulphated CCK to 100 ng led to a considerable prolongation of feeding. CCK-8-S and CCK-8-NS in doses used did not affect the reaction of avoidance in rabbits caused by electrical stimulation of the ventromedial nucleus of the hypothalamus.     

The effect of a novel pentadecapeptide BPC 157 on development of tolerance and physical dependence following repeated administration of diazepam

A novel gastric pentadecapeptide BPC 157 with different beneficial activities and anticonvulsant effect interacting with GABAergic system could improve diazepam efficacy coadministered (10 microg/kg, 10 ng/kg i.p.) with diazepam (5.0 mg/kg i.p.) twice daily for 10 days, since diazepam chronic medication would otherwise predispose for diazepam- tolerance/withdrawal development (shorter latency to convulsion after convulsant). In diazepam chronically treated mice, it attenuated diazepam tolerance (provoked by later acute administration of diazepam together with convulsant) and postponed physical dependence/withdrawal effects (provoked by later administration of isoniazid). In tolerance assay, at 42 h after the end of conditioning regimen, shorter preconvulsive latencies than in healthy (non-diazepam conditioned) mice following isoniazid (800 mg/kg i.p.) (as hallmark of tolerance) were observed if diazepam (5.0 mg/kg i.p.) was again given acutely to mice previously conditioned with diazepam alone (use of picrotoxin 3.0 mg/kg i.p., as convulsant, with acute application of diazepam in previously diazepam conditioned mice did not lead to tolerance hallmark). This was completely avoided in diazepam+BPC 157 10 microg or diazepam+BPC 157 10 ng chronically treated animals. In physical dependence assay (isoniazid challenge assessed at 6, 14, 42 and 72 h after conditioning medication), when compared to diazepam non-conditioned healthy mice, in diazepam conditioned mice residual anticonvulsive activity was not present already at the earliest post-conditioning interval (i.e., not different latency to isoniazid-convulsions), whereas shorter preconvulsive latencies (as physical dependence/withdrawal hallmark) were noted in diazepam conditioned mice following isoniazid challenge at 42 h and at 72 h after end of conditioning treatment. In diazepam+BPC 157 10 microg- conditioned mice, a residual anticonvulsive activity (i.e., longer latency to isoniazid convulsion) was noted at 6 h post-conditioning, whereas shorter preconvulsive latencies appeared only at 72 h-post-conditioning period. In conclusion, taken together these data (lack of tolerance development (tolerance studies), prolonged residual anticonvulsive activity, and postponed physical dependence/withdrawal hallmark in diazepam+BPC 157 chronically treated mice) with common benzodiazepines tolerance/withdrawal knowledge, it could be speculated that BPC 157 acts favoring the natural homeostasis of the GABA receptor complex as well as enhancing the GABAergic transmission, and having a mechanism at least partly different from those involved in diazepam tolerance/withdrawal, it may be likely used in further therapy of diazepam tolerance and withdrawal.     

CCK-resistance in Zucker obese versus lean rats

Obese Zucker rats are less sensitive to the satiety effect of CCK than lean litter mates. The present studies further characterised this CCK resistance. Subcutaneous injection of the CCK agonist caerulein dose-dependently decreased food intake in Zucker obese and lean rats whereas the CCK-B agonist gastrin-17 did not. Caerulein at 4 μg/kg, which resulted in CCK plasma bioactivity slightly above postprandial levels, decreased food intake in lean rats but not in obese rats. The decrease in food intake was also more marked at higher caerulein doses (20–100 μg/kg) in lean versus obese rats. In lean animals the satiety effects of the “near physiological” 4 μg/kg caerulein dose was abolished after blockade of vagal afferents with capsaicin, whereas the effects of higher caerulein doses were not. CCK-stimulated amylase secretion from pancreatic acini and binding capacity of ¹²⁵I- labelled CCK-8 were decreased in obese versus lean rats. The CCK-A antagonist loxiglumide at 20 mg/kg, a dose which abolished the action of all caerulein doses on food intake, failed to alter the food intake either in obese or in lean rats when given without an agonist. The results suggest that the satiety effects of “near physiological” doses of caerulein in lean rats are mediated by vagal afferents whereas pharmacological doses act via non-vagal mechanisms. The differences in CCK's satiety effect between lean and obese rats may be due to differences in CCK-receptor binding and action at peripheral vagal sites. However, the failure of the CCK-A antagonist to increase food intake questions whether any of the effects of exogenous CCK are of physiological relevance.     

Partial reversal of behavioral action of the agonist D-Trp-6-LH-RH by naloxone in mice

The effects of the superactive agonist analog D-Trp-6-LH-RH were investigated in several neuropharmacological tests: inhibition of picrotoxin-induced seizures, open-field behavior, hot-plate and tail-flick tests, assessment of catalepsy and apomorphine-induced cage-climbing. In most tests, D-Trp-6-LH-RH was administered subcutaneously (sc.) at the dose of 100 micrograms/kg. The opiate involvement in the peptide action was checked by using naloxone HCl (NX) in a dose of 1 mg/kg intraperitoneally (ip.), with the exception of the analgesic tests where the dose was 0.5 mg/kg. The analog significantly suppressed the open-field parameters of ambulation, rearing and grooming; except for grooming, these actions were fully antagonized by NX. Similarly, NX pretreatment restored to the control levels the latencies of seizure parameters increased by D-Trp-6-LH-RH. The hot-plate latencies did not change after pretreatment with NX but the opiate antagonist was fully able to antagonize the analgesic effect of the peptide in the tail-flick test. The cataleptogenic effect and the inhibition of apomorphine-induced cage-climbing demonstrated after D-Trp-LH-RH were not antagonized by NX.     

八肽胆囊收缩素对抗阿片肽的中枢性降血压作用

过去的工作已经证明八肽胆囊收缩素(CCK-8)能够对抗阿片肽的镇痛作用,本工作探讨CCK-8是否能够对抗阿片肽的心血管抑制作用。给戊巴比妥钠麻醉大鼠脊髓蛛网膜下腔(ith)注射 CCK-8可以对抗 ith 注射 mu(μ)型阿片受体激动剂[NMePhe~3,D-Pro~4]Morphiceptin(PL017)(5μg)、delta(δ)型受体激动剂[D-Ala~2,D-Leu~5]Enkephalin(DADLE)(25μg)和 Kappa(K)型受体激动剂[N-Me Tyr,N-Me Arg~7,D-Leu~8]Dynorphin 1-8 ethyla-mide(66A-078)(1μg)引起的降低血压和减慢心率作用。在 MAP 的表现上,CCK-8的拮抗作用(10μg及以下剂量)具有量-效关系,并可被 CCK 受体阻断剂丙谷胺(Proglumide)(100μg)翻转。在 HR 的表现上,上述剂量的 CCK-8也显示了一定的拮抗作用,但量-效关系不如 MAP 表现得明显。单纯将 CCK-8或 Proglumide ith 注射,可见大剂量(50μg)CCK-8可以引起明显的降血压作用和短时的降心率作用,小剂量(0.05μg)CCK-8则表现出明显的降心率作用;ith 注射 Proglumide 100μg,30 min 后也表现出减慢心率的作用。以上结果提示:在脊髓水平,一定剂量范围内的 CCK-8能够对抗阿片肽的心血管抑制效应,此对抗作用是通过 CCK 受体实现的。本工作的结果支持关于 CCK-8是一种抗阿片物质的设想。     

Influence of cerebro-spinal fluid of picrotoxin kindled rats on seizure susceptibility and locomotor activity of recipients.

Repeated picrotoxin administration (ip) in subthreshold doses in rats resulted in kindling of generalized seizures. Decrease of locomotor activity in kindled rats occurred in interictal periods. Intra-cerebroventricular microinjection to intact recipients of cerebrospinal fluid (CSF) of kindled but not intact rats or those after acute picrotoxin-induced convulsions, induced a decrease of locomotor activity and severity of acute picrotoxin induced seizures. These effects of CSF were blocked by naloxone pretreatment and were absent after injection of CSF to which protease inhibitors were not added. It is concluded that the release of endogenous opioid peptide substance(s) takes place in CSF of kindled animals which cause the interictal decrease of locomotor activity and may play the role of endogenous anticonvulsive factors controlling epileptic activity induction.     

Effect of cholecystokinin receptor antagonists, MK-329 and L-365,260, on cholecystokinin-induced acid secretion and histidine decarboxylase activity in the rat

To elucidate the regulatory mechanism of acid secretion by cholecystokinin (CCK) in vivo, we compared the effects of CCK and gastrin on acid secretion and histidine decarboxylase (HDC) activity. We also examined the effects of MK-329, a specific antagonist for pancreatic-type CCK receptor, and L-365,260, a specific antagonist for gastrin-type CCK receptor, on the action of CCK. Graded doses of CCK or gastrin were intravenously infused into conscious rats with gastric fistula. Gastrin-17 I infusion up to 10 nmol/kg/h resulted in dose-related increases in acid secretion. CCK-8 infusion also caused an increase in acid secretion. However, it reached a peak with 0.3 nmol/kg/h CCK-8 and attenuated with higher concentrations of CCK-8. This attenuating effect of a higher dose of CCK was reversed by MK-329, but not by L-365,260. Both CCK and gastrin were potent in increasing fundic HDC activity, and the effect of CCK on HDC activity was significantly inhibited by L-365,260, but not by MK-329. Taken together, the present study suggests that CCK and gastrin stimulate histamine formation via a gastrin-type CCK receptor, and the attenuating action of CCK with higher concentrations on acid secretion in vivo is mediated by a pancreatic-type CCK receptor.     

The participation of CCK-8-ergic mechanisms in the regulation of emotional behavior in rodents

Effects of CCK-8 receptor agonists caerulein and pentagastrin and CCK-8 receptor antagonist proglumide on exploratory and locomotor activity of mice and rats were studied. Systemic administration of caerulein (500 ng/kg 1 mcg/kg) decreased significantly the exploratory activity of mice in elevated plus-maze. This anxiogenic-like action of caerulein was attenuated by acute pretreatment with proglumide (1 and 15 mg/kg) but not with diazepam (up to 0.75 mg/kg). Proglumide slightly increased the exploratory activity of rats in plus-maze; on the other hand, caerulein and pentagastrin potently decreased the measures of exploration in this test. Caerulein (10-100 mcg/kg) and proglumide (1 and 15 mg/kg) inhibited 3H-pentagastrin binding in mice brain in in vivo experiments. The data obtained indicate that CCK-8-ergic mechanisms in brain play an important role in the generation of anxiety states in rodents.     

Pharmacological manipulation of sincalide (CCK-8)-induced suppression of feeding

The current study involves an investigation of the possible neurotransmitter systems involved in the ability of exogenously administered sincalide (cholecystokinin octapeptide, CCK-8) to suppress feeding. Male rats previously trained to obtain food either during a daily 3-hr session, or conditioned to obtain food pellets on a fixed-ratio or fixed-interval schedule of reinforcement, were treated IP with CCK-8, following pretreatment with representative drugs of several pharmacological classes. Pretreatment with phenoxybenzamine, tolazoline, yohimbine, morphine, haloperidol or picrotoxin reduced the efficacy of CCK-8. However, pretreatment with naloxone or clonidine potentiated the suppressant action of CCK-8 on feeding. Propranolol, diphenhydramine, cimetidine, atropine, d-amphetamine, fenfluramine or diazepam pretreatment either had no effect or no consistent action in altering the activity of CCK-8. The ability of CCK-8 to suppress feeding was not altered by subacute treatment with the anorectics, d-amphetamine or fenfluramine, using a regimen known to induce tolerance. These data indicate that CCK-8 exerts a different mechanism of action than that of fenfluramine or d-amphetamine, and furthermore, that noradrenergic, dopaminergic, GABAergic or endogenous opioid systems either mediate or can modify the effect of CCK-8 on feeding.     

MK-329 blocks the inhibition of alcohol intake by CCK-8

Peripheral administration of sulfated cholecystokinin octapeptide (CCK-8) potently reduces alcohol intake, preference, and blood levels in rats. MK-329 (L-364,718 or Devazepide) acts at peripheral cholecystokinin (CCK_A) receptors to antagonize CCK-8's physiological and behavioral effects, such as pancreatic stimulation and inhibition of feeding. We determined whether CCK_A receptor blockade would also prevent CCK-8's alcohol satiety effect. Water-deprived female and male rats (n = 7 for each) received randomized combinations of intraperitoneal injections of MK-329 (0, 100, 200, or 400 μg/kg) followed by CCK-8 (0 or 4 μg/kg). Rats were then given access to 5% w/v ethanol for 30 min, followed by 30-min access to water, with food ad lib. MK-329 at all doses significantly (p < 0.05) reduced the suppression of alcohol intake and food intake by CCK-8. MK-329 alone increased alcohol intake at 400 μg/kg, and increased food intake, in females and males at 100 and 200 μg/kg, respectively. We concluded that CCK-8's alcohol and food satiation effects depend on specific, peripheral CCK_A receptors, and satiation of alcohol consumption and drinking-associated feeding reflect an endogenous functional interaction of CCK-8 with CCK_A receptors.     

Effect of diazepam, carbamazepine, sodium valproate and their combinations with vitamin preparations on epileptic activity

The anticonvulsive action of diazepam, carbamazepine, sodium valproate and their combinations with pyridoxal-5-phosphate, nicotinamide, and alpha-tocopherol were investigated in acute experiments on mice with corazole-induced seizures. Diazepam (0.5 mg/kg), carbamazepine (50 mg/kg) and sodium valproate (200 mg/kg) were shown to reduce convulsive intensity and lethality. Vitamins nicotinamide (250 mg/kg), pyridoxal-5-phosphate (10 mg/kg) and alpha-tocopherol (100 mg/kg) potentiated anticonvulsive action of the above antiepileptic drugs. The results of the investigation suggest the efficacy of pathogenetic therapy and give new evidence of the advisability of using vitamins in combination with synthetic anticonvulsive drugs.     

A possible interaction between CCKergic and GABAergic systems in the rat brain

Cholecystokinin sulfated octapeptide (CCK-8S) was given to rats i.p. at single doses of 10 and 100 nmol/kg, respectively. It produced a modification in GABA levels in several areas of the rat brain. After 30 min of injection, the lower dose (10 nmol/kg) increased GABA levels in striatum by 31% (P<0.05). The higher dose (100 nmol/kg) enhanced GABA levels either in hippocampus by 78% (P<0.05) or in frontal cerebral cortex by 81% (P<0.05) and decreased in olfactory bulbs by 57% (P<0.01). Thus, these results show that systemic injection of CCK-8S, produced regional specific changes on GABA levels in brain, and these effects were dose-dependent. Systemic pretreatment with the CCK(B) receptor antagonist, PD 135,158, 1 mg/kg i.p., on the endogenous levels of GABA in certain regions was also studied. The selective CCK(B) receptor antagonist, PD 135,158, did not have an effect per se on the endogenous levels of GABA but prevents the action induced by the neuropeptide. We suggest that the action of CCK may be mediated via a selective action on the CCK(B) receptor subtypes.     

Role of cAMP-Dependent Protein Kinase on Acute Picrotoxin-Induced Seizures

cAMP-dependent protein kinase (PKA) is a major modulator of synaptic transmission likely to be involved in molecular and cellular events leading to epileptogenesis, but little is known about how it affects the onset of acute epileptic seizures. In this study, we determined PKA enzymatic activity in the rat hippocampus during picrotoxin-induced seizures, using H-9 dihydrochloride, a PKA inhibitor, to investigate the in vivo effects of this enzyme on seizures induced by picrotoxin microdialysis in the rat hippocampus. No significant modifications were found in PKA activity during seizures as compared to control rats, but H-9 dihydrochloride microperfusion (100 μM) prevented picrotoxin seizures in 50% of the animals and significantly reduced the mean number of seizures and mean seizure duration. These results suggest that acute picrotoxin-induced seizures occur without an increase in hippocampal PKA activity, but reduced PKA-mediated phosphorylation protects against picrotoxin seizures, probably by increasing the inhibitory potential of GABA_A receptors. The possibility of other targets for H-9 dihydrochloride, such as PKC, PKG or CAMKII, however, cannot be ruled out.