Dopamine receptors in the stellate ganglion of the dog

Effects of fenoldopam, a selective DA1 dopamine receptor agonist, and dipropyl dopamine and propylphenethyl dopamine, preferential DA2 dopamine receptor agonists, on ganglion transmission were studied in pentobarbital-anesthetized, open-chest dogs. Tachycardia induced by electrical stimulation (supramaximal voltage, 0.5 ms duration, 1-2 Hz) of the preganglionic cardio-accelerator nerves was monitored as a measure of ganglionic transmission. Drugs were injected into the costocervical artery (i.a.) close to the arterial supply of the ganglion. Doses required to produce 30-40% inhibition of ganglionic transmission by the i.a. route were 2-8 micrograms for dipropyl dopamine, 4-16 micrograms for propylphenethyl dopamine, and 100 micrograms for fenoldopam. At these doses none of the agonists affected tachycardia induced by electrical stimulation of the postganglionic nerve. Domperidone (5 micrograms/kg i.v.), a selective DA2 dopamine receptor antagonist, markedly antagonized the effects of dipropyl dopamine and propylphenethyl dopamine, but had only minor (and statistically insignificant) effects on the inhibitory effect of fenoldopam. SCH 23390 (5 micrograms/kg i.v.), a selective and potent DA1 antagonist, failed to modify the effects of any of the agonists. In a separate series, infusion of fenoldopam, 20 micrograms/kg per min i.v. for 5-7 min, facilitated postganglionic nerve stimulation and blocked the inhibitory effect of UK 14,304, an alpha 2-adrenoceptor agonist, on the postganglionic nerve. These results confirm and support the presence of DA2 but do not support the presence of the prototypal DA1 dopamine receptor in the mammalian ganglia. Furthermore, the alpha 2-adrenoceptor blocking property of fenoldopam points to the complication of using i.v. administration for studying its ganglionic actions while monitoring the target tissue effects in response to preganglionic nerve stimulation.     

Interaction of CCK with pancreatic acinar cells

Recent studies demonstrate that cholecystokinin-like peptides are widely distributed in the CNS as well as in the peripheral nervous system and gastrointestinal tract. Studies with agonists have demonstrated multiple classes of receptors and recently potent receptor antagonists have been described which will distinguish these classes and should allow a better understanding of the role of CCK in various physiological processes. One of the known peripheral physiological functions of CCK is the stimulation of digestive enzymes from pancreatic acinar cells. In recent years the interaction of CCK with pancreatic acinar cells has been extensively studied and significant advances have been made in understanding its cellular basis of action. Robert Jensen and colleagues report on each of these areas.     

Dopamine released from nerve terminals activates prejunctional dopamine receptors in dog mesenteric arterial vessels.

The fractional release of dopamine and noradrenaline (NA) from the main trunk of the dog mesenteric artery and its proximal branches, when elicited by K+ (52 mM), was measured by high pressure liquid chromatography with electrochemical detection. K+-induced depolarization released both dopamine and NA. For the main trunk of the mesenteric artery, the fractional release of dopamine and NA were of the same order of magnitude, whereas for the proximal branches dopamine fractional release was significantly lower than that of NA. Phentolamine (0.2 microM) significantly increased dopamine and NA release in both segments of the mesenteric artery. However, for the proximal branches the effect of phentolamine on dopamine and NA release was greater than that observed in the main trunk. Sulpiride (1 microM) significantly increased dopamine and NA release in the proximal branches of the mesenteric artery, whereas in the main trunk sulpiride did not increase amine release. In the proximal branches of the mesenteric artery, sulpiride significantly enhanced dopamine and NA fractional release even after it had been augmented by phentolamine. Apomorphine (0.3 microM) significantly reduced dopamine and NA release in both segments of the mesenteric artery under study; this effect was abolished by sulpiride but not by phentolamine. These results suggest that dopamine and NA released during depolarization by K+ activate prejunctional dopamine and alpha-adrenoceptors, respectively, thereby playing a role in the control of transmitter release.     

NO/cGMP pathway is involved in exocrine secretion from rat pancreatic acinar cells

The enzyme responsible for the synthesis of nitric oxide (NO) from L-arginine in mammalian tissues is known as nitric oxide synthase (NOS) (EC.1.14.13.39). In the present study, the role of NO in the regulation of exocrine secretion was investigated in rat pancreatic acinar cells. Treatment of rat pancreatic acinar cells with cholecystokinin-octapeptide (CCK-OP) resulted in an increase in the arginine conversion to citrulline, the amount of NO_x, the release of amylase, and the level of cGMP. Especially, CCK-OP-stimulated increase of arginine to citrulline transformation, the amount of NO_x and cGMP level were completely counteracted by the inhibitor of NOS, N^G-monomethyl-L-arginine (MMA), by contrast, that of amylase release was partially reduced. Furthermore, MMA-induced decrease of NOS activity and amylase release showed dose-dependent pattern. The data on the time course of CCK-OP-induced citrulline formation and cGMP rise indicate that NOS and guanylate cyclase were activated by treatment of CCK-OP. However, the mechanism of agonist-stimulated guanylate cyclase activation in acinar cells remains unknown. Therefore, activation of NOS is one of the early events in receptor-mediated cascade of reactions in pancreatic acinar cells and NO, not completely, but partially mediate pancreatic enzyme exocrine secretion.     

Stimulus-secretion coupling and Ca2+ dynamics in pancreatic acinar cells

1. Unique spatiotemporal dynamics in cytosolic Ca²⁺ concentration, [Ca²⁺]_c, were characterized in various cell types. In pancreatic acinar cells, physiological concentrations of cholecystokinin octapeptide, CCK-8, (<10 pM) induce repetitive [Ca²⁺]_c spikes commonly termed Ca²⁺ oscillation, whereas relatively higher concentrations (30 pM-1 nM) evoke biphasic [Ca²⁺]_c dynamics; a rapid transient peak followed by a sustained increase. Much higher concentrations ( > 1 nM) induce a large transient followed by a steep decay.2. These [Ca²⁺]_c dynamics correspond to secretory responses. Repetitive [Ca²⁺]_c change is attributable to the upstroke of the bell-shaped dose-response relationship and the biphasic change is responsible for the downstroke of the relation (so called high-dose inhibited secretion). The large transient [Ca²⁺]_c increase is associated with morphological changes such as bleb formation.3. Possible interrelation between dose of secretagogues, secretory responses, [Ca²⁺]_c dynamics, IP₃ production, receptor occupation and morphological change will be discussed from both pharmacological and physiological points of view.     

The role of nicotinic receptors in dog pancreatic exocrine secretion.

1 The effects of pentamethonium, an autonomic ganglion blocker, were studied on the exocrine pancreatic secretion of six conscious dogs given intravenous infusions of urecholine, caerulein or pentagastrin on a background of submaximal doses of secretin. 2 Urecholine-induced protein secretion was not affected but both caerulein- and to a smaller extent, pentagastrin-induced protein secretions were depressed by pentamethonium. 3 These results indicate that intravenous caerulein and pentagastrin, but not urecholine, act at least partially via nicotinic receptors. 4 Volume and bicarbonate output were depressed by pentamethonium when stimulated by intravenous caerulein with a background of secretin, but not when stimulated by pentagastrin on a background of secretin. 5 From these data it is suggested that caerulein and pentagastrin may potentiate secretin-stimulated hydrelatic secretion by different mechanisms.     

多器官功能障碍大鼠胰腺腺泡细胞超微结构和功能的改变

目的探讨多器官功能障碍综合征（MODS）时胰腺腺泡细胞超微结构和功能的改变。方法实验地点为福建省立医院动物房、福建省立医院心血管研究所重点实验室、福建省立医院病理科与福建医科大学电镜室。清洁级健康雄性SD大鼠20只,体重180～220g,随机分成2组,即：MODS组和对照组,每组10只。MODS组大鼠腹腔内注射大肠杆菌菌液,同时钳断大鼠左下肢,造成感染复合创伤二次打击MODS大鼠模型。对照组给予等量生理盐水腹腔内注射。48h后检测有两个以上器官或系统功能障碍即为MODS组造模成功。成功造模后,抽血测定其血清胰淀粉酶、脂肪酶,两组大鼠资料的比较用成组t检验。抽血后取标本电镜下观察两组大鼠胰腺腺泡细胞超微结构,免疫组化分析胰腺腺泡细胞Bax蛋白表达。结果和对照组比较,MODS组大鼠胰淀粉酶、脂肪酶及Bax蛋白表达明显高于正常对照组（P〈0.01）。胰腺腺泡细胞出现空泡变性,少数细胞坏死,及出现线粒体肿胀,粗面内质网扩张等一系列超微结构病理改变。结论 MODS大鼠可出现胰腺损害。     

Emodin attenuates cell injury and inflammation in pancreatic acinar AR42J cells

The study was intended to investigate the protective effects of emodin against cell injury and inflammation in AR42J cells. We determined trypsin and lipase activity, intracellular ROS and MMP using specific assay kits. The related protein expression and TNF-α and IL-6 in the medium were assayed by Western blot and ELISA kits. Results showed that emodin could protect AR42J cells against cell injury caused by cerulein and lipopolysaccharide which were possibly associated with inhibition of mitochondrial damage, ROS production, and then significantly inhibited ROS-mediated pathway, and ameliorated pancreatic cells injury by depleting the levels of inflammatory cytokines.     

脂多糖对离体大鼠胰腺腺泡细胞钙稳态的影响

目的:研究脂多糖(LPS)对胰腺腺泡细胞钙稳态的影响,及胞浆内钙超载时钙离子的来源,以探讨LPS致胰腺腺泡细胞损伤的机制。方法:胶原酶法分离胰腺腺泡细胞,Fluo-3/AM负载后,在无钙或含生理钙离子浓度的培养液中加入不同浓度LPS,采用激光共聚焦显微镜观察单个胰腺腺泡细胞[Ca~(2+)]_i。另外采用MTT法检测LPS作用的不同时间点胰腺腺泡细胞的活性。结果:LPS可致胰腺腺泡细胞损伤、细胞内[Ca~(2+)]_i显著升高,且呈浓度依赖性(P<0.05):在含1mmol/L依他酸的培养液中,LPS致腺泡细胞损伤程度明显减轻(P<0.05)、仅引起胰腺腺泡细胞[Ca~(2+)]_i缓慢、微弱的升高,而再次恢复培养液中Ca~(2+)至生理浓度,引发快速、幅度更高而持久的[Ca~(2+)]_i变化。腺泡细胞内[Ca~(2+)]_i的变化明显先于腺泡细胞的损伤。结论:LPS导致胰腺腺泡细胞内钙超载,主要源于胞外Ca~(2+)内流。钙超载作为早期的病理事件参与腺泡细胞损伤的发生,钙稳态失衡是LPS致胰腺细胞损伤的主要因素之一。     

Dopamine receptors in human platelets

The expression of dopamine receptors by human platelets was investigated by Western blot analysis and immunocytochemical techniques using antibodies raised against dopamine D1-D5 receptor protein. The influence of dopamine D1-like and D2-like receptor agonists on adrenaline-induced platelet aggregation was also investigated. Western blot analysis revealed that platelet membranes bind anti-dopamine D3 or D5 receptor protein antibodies, but not anti-D1, D2 or D4 receptor protein antibodies. Cytospin centrifuged human platelets exposed to anti-dopamine D3 or D5 receptor protein antibodies developed a specific immune staining, whereas no positive staining was noticeable in platelets exposed to other antibodies tested. Both the D1-like receptor agonist 1-phenyl2,3,4,5-tetrahydro-(1H)-3-benzazepine-7,8-diol hydrochloride (SKF 38393) and the D2-like receptor agonist 7-hydroxy-N,N-di-n-propyl-2-aminotetralin (7-OH-DPAT) dose-dependently inhibited adrenaline-induced platelet aggregation. These effects were decreased respectively by the D-like and D2-like receptor antagonists R(+)-2,3,4,5-tetrahydro-3-methyl-5-phenyl-1H-3-benzazepin-7-ol hydrochloride (SCH 23390) and (-)sulpiride. The above findings indicate that human platelets express dopamine D3 and D5 receptors probably involved in the regulation of platelet function.     

Dopamine D4 receptors

Initial investigations on dopamine D4 receptors generated much interest in the role of this receptor in schizophrenia and other aspects of human behavior, as well as new opportunities for novel therapeutics. However, attempts to treat patients suffering from schizophrenia with dopamine D4 agents have failed to yield satisfactory results so far. An examination of the dopamine D4 literature shows that contrasting and conflicting data seemed to be the norm in this field of research. This paper reviews the literature on the dopamine D4 receptor and discusses many of the associated methodological problems that might have contributed to the paradoxical findings.     

Cholecystokinin activates a variety of intracellular signal transduction mechanisms in rodent pancreatic acinar cells

Cholecystokinin (CCK) acting through its G protein-coupled receptor is now known to activate a variety of intracellular signaling mechanisms and thereby regulate a complex array of cellular functions in pancreatic acinar cells. The best studied mechanism is the coupling through heterotrimeric G proteins of the Gq family to activate a phospholipase C leading to an increase in inositol trisphosphate and release of intracellular Ca2+. This pathway along with protein kinase C activation in response to the increase in diacylglycerol stimulates the secretion of digestive enzymes by the process of exocytosis. CCK also activates signaling pathways in acini more related to other processes. The three mitogen activated protein kinase cascades leading to ERKs, JNKs and p38 MAPK are all activated by CCK. CCK activates the ERK cascade by PKC activation of Raf which in turn activates MEK and ERKs. JNKs are activated by a distinct mechanism which requires higher concentrations of CCK. Both ERKs and JNKs are presumed to regulate gene expression. CCK activation of p38 MAPK also plays a role in regulating the actin cytoskeleton through phosphorylation of the small heat shock protein HSP27. The PI3K-PKB-mTOR pathway is activated by CCK and plays a major role in regulating protein synthesis at the translational level. This includes both activation of p70 S6K leading to phosphorylation of ribosomal protein S6 and the phosphorylation of the binding protein for initiation factor 4E leading to formation of the mRNA cap binding complex. Other signaling pathways activated by CCK receptors include NF-kappaB and a variety of tyrosine kinases. Further work is needed to understand how CCK receptors activate most of the above pathways and to better understand the biological events regulated by these diverse signaling pathways.     

Dopamine receptors in human basilar arteries

After phenoxybenzamine (10(-5) M), pretreatment, and in the presence of propranolol (10(-6) M) and indomethacin (2.8 X 10(-6) M), dopamine caused a marked concentration-dependent relaxation of isolated strips of human basilar artery contracted with PGF2 alpha. This effect was mimicked by apomorphine, 6,7-ADTN and SK&F 38393, but N,N-diethyl dopamine, N,N-di-n-propyl-dopamine and 5,6-ADTN caused only slight relaxation. (+)-Butaclamol, cis-alpha-flupenthixol, fluphenazine and haloperidol competitively antagonised the relaxant effects of dopamine, but sulpiride was ineffective in concentrations as high as 1.3 X 10(-4) M. These findings show that the dopamine receptors in the human basilar artery closely resemble those in the smooth muscle of the rabbit isolated mesenteric and splenic arteries, and the dog renal and mesenteric arteries in vivo, but differ from those located presynaptically on sympathetic nerve terminals.     

Down-regulation of bombesin binding to guinea-pig pancreatic acinar cells during homologous desensitization.

1. [125I]-Tyr4-bombesin exhibited saturable binding to pancreatic acinar cells. 2. Preincubation of cells at 37 degrees C with 0.03 nM-1 microM-bombesin for 10 min followed by acid or neutral washes reduced subsequent binding of [125I]-Tyr4-bombesin in a concentration-dependent manner by up to 90%. 3. In cell suspensions, bombesin raised the cytoplasmic Ca2+ concentration ([Ca2+]i) and the [Ca2+]i response was reduced by pre-exposure to the agonist. 4. In individual superfused cells, bombesin at 1 nM normally caused a large [Ca2+]i transient followed by sustained [Ca2+]i oscillations. The cells exhibited a variable degree of desensitization when subsequently exposed to higher agonist concentrations. 5. Preincubation with bombesin for 10 min caused a concentration-related reduction of subsequent amylase responses to bombesin. 6. Down-regulation of receptor binding is a rapid phenomenon during bombesin exposure explaining, at least partially, tachyphylaxis of [Ca2+]i and amylase responses.     

Role of regulators of g-protein signaling 4 in ca signaling in mouse pancreatic acinar cells

Regulators of G-protein signaling (RGS) proteins are regulators of Ca(2+) signaling that accelerate the GTPase activity of the G-protein α-subunit. RGS1, RGS2, RGS4, and RGS16 are expressed in the pancreas, and RGS2 regulates G-protein coupled receptor (GPCR)-induced Ca(2+) oscillations. However, the role of RGS4 in Ca(2+) signaling in pancreatic acinar cells is unknown. In this study, we investigated the mechanism of GPCR-induced Ca(2+) signaling in pancreatic acinar cells derived from RGS4(-/-) mice. RGS4(-/-) acinar cells showed an enhanced stimulus intensity response to a muscarinic receptor agonist in pancreatic acinar cells. Moreover, deletion of RGS4 increased the frequency of Ca(2+) oscillations. RGS4(-/-) cells also showed increased expression of sarco/endoplasmic reticulum Ca(2+) ATPase type 2. However, there were no significant alterations, such as Ca(2+) signaling in treated high dose of agonist and its related amylase secretion activity, in acinar cells from RGS4(-/-) mice. These results indicate that RGS4 protein regulates Ca(2+) signaling in mouse pancreatic acinar cells.     

Congo red modulates ACh-induced Ca2+ oscillations in single pancreatic acinar cells of mice

Aim:

Congo red, a secondary diazo dye, is usually used as an indicator for the presence of amyloid fibrils. Recent studies show that congo red exerts neuroprotective effects in a variety of models of neurodegenerative diseases. However, its pharmacological profile remains unknown. In this study, we investigated the effects of congo red on ACh-induced Ca²⁺ oscillations in mouse pancreatic acinar cells in vitro.

Methods:

Acutely dissociated pancreatic acinar cells of mice were prepared. A U-tube drug application system was used to deliver drugs into the bath. Intracellular Ca²⁺ oscillations were monitored by whole-cell recording of Ca²⁺-activated Cl⁻ currents and by using confocal Ca²⁺ imaging. For intracellular drug application, the drug was added in pipette solution and diffused into cell after the whole-cell configuration was established.

Results:

Bath application of ACh (10 nmol/L) induced typical Ca²⁺ oscillations in dissociated pancreatic acinar cells. Addition of congo red (1, 10, 100 μmol/L) dose-dependently enhanced Ach-induced Ca²⁺ oscillations, but congo red alone did not induce any detectable response. Furthermore, this enhancement depended on the concentrations of ACh: congo red markedly enhanced the Ca²⁺ oscillations induced by ACh (10–30 nmol/L), but did not alter the Ca²⁺ oscillations induced by ACh (100–10000 nmol/L). Congo red also enhanced the Ca²⁺ oscillations induced by bath application of IP₃ (30 μmol/L). Intracellular application of congo red failed to alter ACh-induced Ca²⁺ oscillations.

Conclusion:

Congo red significantly modulates intracellular Ca²⁺ signaling in pancreatic acinar cells, and this pharmacological effect should be fully considered when developing congo red as a novel therapeutic drug.