Cholecystokinin-like Peptides Research Articles

Sulphated cholecystokinin octapeptide inhibits ethanol consumption in the rat

Twenty-three hr water deprived rats received access to 5% ethanol solution for 30 min daily. Intraperitoneal injection of sulphated cholecystokinin octapeptide (CCK-8, 2.0–16.0 μg/kg) significantly inhibited ethanol consumption, but injection of desulphated CCK-8 did not affect ethanol consumption. These results confirm a previous report that CCK-8 suppresses ethanol intake, and indicate that inhibition of ethanol consumption by CCK-8 depends on sulphation of its tyrosyl residue. This chemical specificity of action of CCK-8 is also characteristic of other known behavioral and physiological effects of peripherally-administered cholecystokinin-like peptides.

Effects of acetylcholine and caerulein on 86Rb + efflux in the mouse pancreas. Evidence for a sodium-potassium-chloride cotransport system

The effect of acetylcholine and the cholecystokinin-like peptide, caerulein on the fractional efflux of 86Rb + from preloaded isolated segments of mouse pancreas were studied. Both secretagogues evoked a marked transient increase in 86Rb + efflux. The removal of Ca 2+ from the superfusing medium and addition of 10 −4 M EGTA, markedly reduced, but did not abolish the responses to either acetylcholine or caerulein. Furosemide ( 10 −5−10 −3 M ) or piretanide (10 −4 M) reduced the basal efflux and inhibited the secretagogue-elicited responses. Stimulus-induced 86Rb + outflow was abolished when the Cl − component of the superfusing solution was replaced by either NO 3 −, SO 4 2− or I − but not in case of replacement by Br −, When Na + was replaced with either Li + or choline + both acetylcholine and caerulein failed to elicit any detectable increase in 86Rb + outflow. However, when Tris + was substituted for Na +, acetylcholine caused a moderate increase in 86Rb + efflux which was abolished by either furosemide (10 −4 M) or chloride depletion (nitrate substitution). The removal of extracellular K + or pretreatment with 10 −3 M ouabain had little effect on secretagogue-evoked 86Rb + efflux. These results indicate the presence of a diuretic-sensitive Na +-K +-Cl − cotransport system in the mouse pancreatic acinar cell membrane.

Immunochemical characterization of gastrinlike and cholecystokininlike peptides released in dogs in response to a peptone meal

This study was designed to examine and compare the nature of gastrinlike and cholecystokininlike peptides released into the portal and peripheral venous circulation in response to a peptone meal. Six dogs were prepared with portal venous catheters and gastric fistulas. Portal and peripheral venous sera were obtained before and after gastric infusion of a 10% peptone meal. Serum levels of gastrin and cholecystokinin immunoreactive peptides were determined by radioimmunoassay using two distinct peptide region-specific antibody preparations. These separate antibody preparations demonstrated specificity for (a) C-terminal tetradecapeptide gastrin (4–17hG17), heptadecapeptide gastrin (G17), and big gastrin (G34) (gastrin antibody); and (b) all biologically active forms of gastrin and cholecystokinin (gastrin-cholecystokinin antibody). Using the antibody preparation with specificity for gastrin and not cholecystokinin, the mean basal immunoreactive gastrin from portal (8.33 ± 2.4 fmol/ml, mean ± SEM) and from peripheral (6.19 ± 0.9 fmol/ml) venous sera both increased after peptone infusion, with an early peak (2 min in portal and 4 min in peripheral serum) and a second peak at 30 min in both circulations. Measurements using antibodies with specificity for both cholecystokinin and gastrin yielded strikingly different results. The portal venous serum peptide concentration (49 ± 10 fmol/ml) increased sharply within 30 s after peptone infusion to a single peak at 2 min (139 ± 37 fmol/ml). The basal peripheral venous serum peptide concentration (43 ± 8.8 fmol/ml) increased more gradually to a single peak at 8 min (78 ± 14 fmol/ml). Studies with Sephadex (G-50 superfine) gel chromatography indicated that gastrin released in response to the peptone meal was primarily G17. However, of the peptides released in response to the peptone meal that were recognized by the gastrin-cholecystokinin antibody, >80% were shown to be distinct from gastrin. Gel chromatographic studies demonstrated that peptone meal-stimulated immunoreactive cholecystokinin release consisted of two major peaks, eluting in positions identical to those of intact cholecystokinin (CCK33) and the octapeptide of cholecystokinin (CCK8).

The Effects of Ceruletide in Schizophrenia

Eight neuroleptic-resistant schizophrenic patients were treated with ceruletide diethylamine, a cholecystokininlike peptide, in a placebo-controlled, double-blind, cross-over study. Ceruletide or placebo was administered intramuscularly twice a day for four consecutive days while patients received a constant dose of fluphenazine hydrochloride. Cholecystokinin octapeptide was also administered to four different schizophrenic patients in a double-blind, cross-over study. Cholecystokinin or placebo was administered as a slow intravenous infusion daily for four days. There were no changes in either the positive or negative symptoms of schizophrenia between the periods of placebo, ceruletide, or cholecystokinin administration. Furthermore, there was no tendency for the patients' conditions to either improve or worsen during the course of ceruletide or cholecystokinin treatment. In contrast to previous reports from uncontrolled studies, cholecystokininlike peptides appear to be devoid of antipsychotic properties when administered parenterally.

Dopamine and cholecystokinin immunoreactive neurones in mesencephalic grafts reinnervating the neostriatum: Evidence for selective growth regulation

Pieces of embryonic mesencephalic tissue rich in dopamine and cholecystokinin immunoreactive neurones were grafted to the dorsal surface of the caudate-putamen of adult host rats subjected to unilateral dopamine depleting lesions. After 3 months, neuronal survival in the graft and fibre outgrowth into the host brain were studied by tyrosine hydroxylase and cholecystokinin immunohistochemistry, both in serial sections and by elution and restaining of the same sections. Both dopamine- and cholecystokinin-containing neurones as well as neurones containing both compounds survived the transplantation process. The ratio of neurones in which dopamine and cholecystokinin-like immunorcactivity occurred independently and in coexistence was similar in the grafts to that seen in the intact ventral mesencephalon. This suggests that the grafted cells maintain and express at least some of their normal chemical characteristics in the ectopic cortical location. Only those fibres which contained tyrosine hydroxylase but apparently lacked the cholecystokinin-like peptide showed extensive reinnervation of the host neostriatum. The cholecystokinin-positive libres were found in a narrow zone immediately adjoining the graft. These results indicate that the dopaminergic reinnervation of the denervated neostriatum is preferentially carried out by the population of grafted mesencephalic dopamine neurones apparently lacking the cholecystokinin-like peptide. This suggests the presence of growth regulating mechanisms in the denervated neostriatum which selectively favour the ingrowth of fibres from the appropriate dopaminergic neuronal subset. The transplantation technique may therefore provide a powerful tool for the study of neurone target interactions in the establishment of neuronal connections, and of the possible role of peptidergic coexistence in the development and organization of monoaminergic pathways and their innervation patterns.

Effect of cholecystokinin-octapeptide on dopamine release from slices of cat caudate nucleus

The cat caudate nucleus has been reported to possess a rich and fairly even distribution of nerve endings, containing both dopamine- and cholecystokinin-like peptides. In this study, the effect of cholecystokinin-octapeptide (CCK-8) on basal and electrically evoked tritium outflow from slices of cat caudate nucleus previously labeled with [3H]dopamine was examined. Evoked tritium outflow from slices of cat caudate nucleus was Ca2+ dependent and abolished by tetrodotoxin, suggesting that it reflects action potential-induced [3H]dopamine release. In the presence of bovine serum albumin and bacitracin, the sulfated but not the unsulfated form of CCK-8 inhibited both basal and electrically evoked tritium outflow from slices of cat caudate nucleus at very low concentrations. CCK-8 sulfate was efficient in causing this effect in concentrations down to 10(-14) M, and the maximum effect was obtained with 10(-11) M. In contrast, without bovine serum albumin and bacitracin, no inhibitory effect of CCK-8 sulfate was seen, but instead, a marked enhancement of tritium outflow at 10(-7) M was observed. The findings suggest that CCK-8 sulfate in dopamine/CCK coexistence regions is involved in regulating dopamine release.

Neuroendocrine responses to stimulation of the vagus nerves in bursts in conscious calves.

Effects of stimulation of the peripheral ends of the vagus nerves below the heart at 4 Hz continuously, and at 40 Hz for 1 s at 10 s intervals, have been compared in conscious calves below behavioural threshold. Neither pattern of stimulation caused any significant change in mean aortic blood pressure or heart rate but both invariably produced a substantial increase in the flow of intestinal lymph. Each form of stimulation provoked release of glucagon, insulin and pancreatic polypeptide from the pancreas and produced a small but significant rise in mean arterial plasma glucose concentration. The release of gastric inhibitory peptide- and bombesin-like molecules from the gastrointestinal tract was not affected by vagal stimulation whereas release of vasoactive intestinal peptide was observed in response to both patterns of vagal stimulation. Evidence was obtained to suggest that gastrin-like peptides are preferentially released into the bloodstream whereas cholecystokinin-like peptides are not. Vagal stimulation releases somatostatin from the gastrointestinal tract but discontinuous stimulation seems to inhibit the release of somatostatin into the general circulation. The results that have been obtained, employing this particular protocol, suggest that the pattern of the stimulus that is applied to the vagal splanchnic innervation has relatively little effect on neuroendocrine response in this species.

Facilitation of GABA release by cholecystokinin and caerulein in rat cerebral cortex

Cholecystokinin octapeptide and its analogue, caerulein, facilitated the K +-evoked release of 14C-GABA from tissue slices of rat parietal cortex. The effect of caerulein was maximal at 1 nM where an enhancement of 36% was produced. Cholecystokinin octapeptide gave rise to a similar maximal enhancement (29%), but was two orders of magnitude less potent. The enhancement of 14C-GABA release by caerulein was reversed by proglumide, a putative competitive antagonist at the cholecystokinin receptor. The possibility that the cholecystokinin-induced facilitation of GABA release in the cortex is involved in the anticonvulsant properties of cholecystokinin-like peptides is discussed.

Cholecystokinin-like peptides potentiate apomorphine-induced inhibition of dopamine neurons

Cholecystokinin-like peptides potentiate apomorphine-induced inhibition of dopamine neurons

Stimulation of pepsinogen release from isolated gastric glands by cholecystokininlike peptides

Gastric glands isolated from rabbit stomach were employed to study the regulation of pepsinogen secretion by peptide hormones. Cholecystokinin octapeptide (CCK-OP) stimulated pepsinogen secretion with an ED50 of about 1 nM. Caerulein was as effective as CCK-OP but less potent (ED50, 10 nM). Gastrin (HG-17) was found to be a weak stimulus, being only about 20% as effective as CCK-OP or caerulein. Sulfation of CCK-OP and caerulein was found to be important for potency but did not alter efficacy. Peptide stimulation of pepsinogen secretion was unaffected by cimetidine, atropine, or propranolol. Combinations of peptides resulted in less-than-additive responses, as did the combination of peptides with carbachol. In contrast, combination of peptides with isoproterenol resulted in additive responses. Accumulation of the weak base aminopyrine was used to measure acid formation by the gastric glands. The peptides gastrin, CCK-OP, and caerulein were found to be equally effective in stimulating acid formation. The peptide stimulation of pepsinogen secretion was inhibited by dibutyryl cGMP, whereas stimulation of acid formation was not inhibited by the cyclic nucleotide. The results indicate that gastric glands contain at least two peptide receptors distinguishable by sensitivity to dibutyryl cGMP. The peptide receptor associated with pepsinogen secretion appears to be selective for CCK relative to gastrin.

Effect of a Cholecystokinin-like Peptide on Some Neuronal Activities in Rat Brain

Effect of a Cholecystokinin-like Peptide on Some Neuronal Activities in Rat Brain

Effect of Cholecystokinin-Octapeptide, Caerulein, and Pentagastrin on Epithelial Cell Proliferation in the Murine Gallbladder

The growth-promoting effect of cholecystokinin-octapeptide, caerulein, and pentagastrin on gallbladder mucosa was investigated in mice. The acute effects on deoxyribonucleic acid synthesis activity was explored with a [3H]thymidine pulse and autoradiography after subcutaneous injection of the polypeptides. The administration of a supramaximal dose of caerulein or of cholecystokinin-octapeptide induced a significant increase (p less than 0.01) in the nuclear uptake of [3H]thymidine by the gallbladder epithelial cells. The injection of pentagastrin had no effect. Implantation of osmotic minipumps was used for the chronic administration of submaximal doses of caerulein or pentagastrin. Using successive [3H]thymidine pulses and autoradiography, increases in labeling (p less than 0.01) and mitotic indices (p less than 0.05) were observed in the caerulein group, whereas pentagastrin had no effect. In addition, a morphometric method was used to determine the total epithelial cell number in the entire gallbladder after chronic administration of the peptides. A significant (p less than 0.01) epithelial cell hyperplasia was observed to occur after caerulein administration whereas pentagastrin given by the same route had no effect on the epithelial cell population. These data indicate that cholecystokininlike peptides promote epithelial growth in the gallbladder of mice.

Chronic blockade of sciatic nerve transmission by tetrodotoxin does not produce central changes in the dorsal horn of the spinal cord of the rat

Rat sciatic nerves were treated with tetrodotoxin (TTX) for 4–10 days, by implanting a glass capillary tube filled with TTX into the nerve through the epineurium. Following this treatment the somatotopic organization of receptive fields in the L4 dorsal horn cells was mapped. This was done by making a series of microelectrode tracks through the medial L4 dorsal horn, an area of cord normally responding only to foot stimulation. The map was normal in animals treated with TTX. Dorsal horn levels of fluoride-resistant acid phosphatase, substance P, somatostatin, cholecystokinin-like peptide, neurotensin and neurophysin were also normal as assessed from density of staining. These results are discussed in the light of the positive changes that are seen following chronic sciatic nerve section.

Substance P and cholecystokinin-like peptides in Helix neurons and cholecystokinin and serotonin in a giant neuron.

Immunohistochemical procedures revealed that the serotonin-containing cell situated in each metacerebral ganglion of the snail Helix also contains a cholecystokinin-like peptide, but is devoid of substance P. In radioimmunoassays the cholecystokinin-like material showed similarities to the carboxy terminal regions of mammalian cholecystokinin and gastrin. Since much is known about the morphology and synaptic connections of this serotonin neuron, the role of the cholecystokinin-like peptides can now be investigated by neurophysiological methods, thus opening the way to discovering whether a neuron can use more than one transmitter.

Immunohistochemical evidence for cholecystokinin-like peptides in neuronal cell bodies of the rat spinal cord.

Cholecystokinin-like immunoreactivity has been demonstrated by radioimmunoassay and immunocytochemistry in the spinal cord of various mammals, in particular in nerve fibers of the superficial layers of the posterior column, but had not been detected in neuronal cell bodies. We report immunohistochemical evidence for the presence of a group of cholecystokinin-containing neuronal cell bodies in the lumbar spinal cord of the rat. This group of cells is only visualized after direct injection of colchicine into the spinal cord and is located near the central canal in the intermedio-medial nucleus of area X of Rexed.

Distribution of substance P in brain and periphery and its possible role as a co-transmitter.

Substance P is widely distributed in the nervous system. In brain and spinal cord it may act as a transmitter, for example at the central branches of primary sensory neurons. It may also be released from the sensory nerve endings and is thought to be involved in antidromic vasodilatation and in synaptic transmission in autonomic ganglia. In some central neurons substance P is stored together with 5-hydroxytryptamine and thyrotropin-releasing hormone. These neurons project to the ventral horn of the spinal cord, amongst other places. In another system substance P coexists with a cholecystokinin-like peptide. These neurons are localized in the periaqueductal central grey matter and also project to the spinal cord. Finally, injection of a substance P antagonist into the ventral mesencephalon causes marked morphological changes in neurons that contain dopamine, substance P and gamma-aminobutyric acid (GABA).

Peptide-monoamine coexistence: Studies of the actions of cholecystokinin-like peptide on the electrical activity of midbrain dopamine neurons

Recent studies have shown that a cholecystokinin-like peptide coexists in a subpopulation of midbrain dopamine-containing neurons. Using extracellular single unit recording techniques we have found that this peptide increases the activity of some, but not all, dopaminergic neurons (identified on the basis of their electrophysiological features). The responsive neurons were found exclusively in areas which were subsequently shown, using immunocytochemical techniques, to contain both cholecystokinin and the enzyme marker for dopaminergic neurons, tyrosine hydroxylase. When administered intravenously, cholecystokinin increased the firing rate of dopaminergic neurons lying in cholecystokinin-rich areas of the substantia nigra. Cells in cholecystokinin-rich ventral tegmental areas showed more variable responses to comparable doses of the peptide. Iontophoretically-applied cholecystokinin consistently activated dopaminergic cells in the substantia nigra and ventral tegmental area, increasing their firing rate and their bursting activity. Cholecystokinin increased the firing rate of some of those neurons to the extent that they were driven into apparent depolarization inactivation. Furthermore, iontophoresis of cholecystokinin resulted in an activation of a population of normally quiescent dopaminergic cells. These results are discussed in light of a possible functional role of cholecystokinin-like peptides in the brain dopaminergic systems. It is suggested that cells in the dopamine-rich areas of the mesencephalon can be characterized both on the basis of their content of peptide and/or catecholamine and of their responsiveness to cholecystokinin-like peptides.

Extraction and immunochemical characterization of cholecystokinin-like peptides from pig and rat brain.

Two major classes of immunoreactive cholecystokinin peptides (iCCK) have been identified in rat and pig brains: (i) large basic peptides (big iCCK) resembling the 33-amino acid porcine cholecystokinin (pCCK33) in size and charge; (ii) small acidic peptides (small iCCK) resembling the COOH-terminal fragments of CCK. Boiling 0.1 M HCl maximally extracts big iCCK; boiling 0.1 M NaOH maximally extracts small iCCK. The differences in hormonal forms removed by these extractants are not likely to be due to enzymatic conversion during the extraction procedures. Fractionation on Sephadex G-50 and starch gel electrophoresis combined with radioimmunoassay using three antisera of different specificities--(i) directed towards the NH2 terminus of pCCK33, (ii) produced by immunization with COOH-terminal fragment CCK8, (iii) produced by immunization with COOH-terminal fragment CCK4--are consistent with the hypothesis that a major fraction of big iCCK may represent intact cholecystokinin with a COOH-terminal extension, as has recently been suggested for gastrin, a molecule having a COOH-terminal pentapeptide identical with that of cholecystokinin.

The release of pancreatic polypeptide by CCK-octapeptide and some analogues in the dog

In dogs with gastric and pancreatic Thomas fistulas the effect of different cholecystokinin-like peptides upon pancreatic polypeptide (PP) release was studied in three ways: (a) Plasma PP concentrations were determined by radioimmunoassay in response to 135 pmol/kg/h of the synthetic C-terminal octapeptide of cholecystokinin (CCK-OP) (A), of three of its analogues (B, C, D) where methionine has been replaced by methoxinine, and in response to 45 pmol/kg/h of caerulein. The greatest rise in plasma PP concentration expressed as ΔPP was achieved with caerulein (327 ± 37 pM), when taking into account the threefold smaller dose used, followed by CCK-OP (536 ± 67 pM) and analogues B (343 ± 51 pM), C (87 ± 46 pM), and D (32 ± 15 pM). The order of potency with respect to stimulation of exocrine pancreatic secretion was the same: E and A precede B, C, and D. ΔPP correlated linearly with the pancreatic protein output (r = 0.98, p < 0.01). (b) CCK-OP was infused in four doses of 35, 70, 135, and 270 pmol/kg/h, and plasma PP concentrations and exocrine pancreatic secretion were monitored. The correlation between pancreatic protein output and ΔPP was very close (r = 0.98, p < 0.01). (c) Atropine sulfate (0.1 mg/kg, i.v.) reduced the PP response to the 135 pmol/kg/h dose of CCK-OP by 61%. We conclude from this that CCK-OP and related peptides do release PP and that their effect on exocrine pancreatic secretion is closely correlated with their PP-releasing capacity. The PP release may therefore be used as an indicator of the CCK-like activity of CCK fragments and analogues. CCK-OP may well represent one of the humoral stimulatory factors contributing to the release of PP, but this action appears to depend on a cholinergic background.

Degradation of cholecystokinin-like peptides by a crude rat brain synaptosomal fraction: A study by high pressure liquid chromatography

Degradation of CCK-8, CCK-4, and related peptides by a crude synaptosomal fraction of rat brain was investigated by monitoring the tryptophan fluorescence of reaction products after HPLC fractionation. At 20°C, the half disappearance time was 52 min for CCK-8, 35 min for unsulphated CCK-8, 20 min for unsulphated CCK-7, 6 min for Tyr(SO 3H)-Trp-Met-Asp-Phe-NH 2, and 3 min only for CCK-4. Caerulein was much more resistant than CCK-8,and Boc-CCK-4 and Aoc-CCK-4 remained stable for at least 3 h. The apparent K m for CCK-8 and CCK-4 was 40 μM and maximal activity on CCK-8 was observed at pH 7.0. Zn 2+ was strongly inhibitory. The protease inhibitors puromycin and bacitracin, the metal chelator 1,10-phenanthroline, and the sulphydryl blocking agents N-ethylmaleimide and p-chloromercuribenzoate greatly reduced the release of tryptophan from CCK-8. Puromycin inhibition of CCK-8 degradation provoked the accumulation of a CCK-7-like peptide, and that of CCK-4 degradation was of a competitive type ( K i = 2 μM). The CCK-8 degrading activity of brain synaptosomes was present in the cytosol as well as in synaptic membranes.