Modulation of the adaptive response to stress by brain activation of selective somatostatin receptor subtypes

Somatostatin-14 was discovered in 1973 in the hypothalamus as a peptide inhibiting growth hormone release. Somatostatin interacts with five receptor subtypes (sst₁₋₅) which are widely distributed in the brain with a distinct, but overlapping, expression pattern. During the last few years, the development of highly selective peptide agonists and antagonists provided new insight to characterize the role of somatostatin receptor subtypes in the pleiotropic actions of somatostatin. Recent evidence in rodents indicates that the activation of selective somatostatin receptor subtypes in the brain blunts stress-corticotropin-releasing factor (CRF) related ACTH release (sst_2/5), sympathetic-adrenal activaton (sst₅), stimulation of colonic motility (sst₁), delayed gastric emptying (sst₅), suppression of food intake (sst₂) and the anxiogenic-like (sst₂) response. These findings suggest that brain somatostatin signaling pathways may play an important role in dampening CRF-mediated endocrine, sympathetic, behavioral and visceral responses to stress.     

Decahydroisoquinoline derivatives as novel non-peptidic,potent and subtype-selective somatostatin sst3 receptor antagonists

Starting from non-peptidic sst₁-selective somatostatin receptor antagonists, first compounds with mixed sst₁/sst₃ affinity were identified by directed structural modifications. Systematic optimization of these initial leads afforded novel, enantiomerically pure, highly potent and sst₃-subtype selective somatostatin antagonists based on a (4S,4aS,8aR)-decahydroisoquinoline-4-carboxylic acid core moiety. These compounds can efficiently be synthesized and show promising PK properties in rodents.     

Expression of Somatostatin and Somatostatin Receptor Subtypes 1�C5 in Human Normal and Diseased Kidney

Somatostatin mediates inhibitory functions through five G protein–coupled somatostatin receptors (sst_1–5). We used immunohistochemistry, immunofluorescence, and RT-PCR to determine the presence of somatostatin receptors sst₁, sst_2A, sst_2B, sst₃, sst₄, and sst₅ in normal and IgA nephropathy human kidney. All somatostatin receptors were detected in the thin tubules (distal convoluted tubules and loops of Henle) and thick tubules (proximal convoluted tubules) in the tissue sections from nephrectomy and biopsy samples. Immunopositive sst₁ and sst₄ staining was more condensed in the cytoplasm of tubular epithelial cells. In normal kidney tissue sections, podocytes and mesangial cells in the glomeruli stained for sst₁, sst_2B, sst₄ and sst₅, and stained weakly for sst₃. In IgA kidney tissue, the expression of somatostatin receptors was significantly increased with particular immmunopositive staining for sst₁, sst_2B, sst₄, and sst₅ within glomeruli. In the epithelial cells, the staining for sst_2B and sst₄ in proximal tubules and sst₁, sst_2B, and sst₅ in distal tubules was increased. The mRNA expression of sst_1–5 was also detected by RT-PCR. Somatostatin and all five receptor subtypes were ubiquitously distributed in normal kidney and IgA nephropathy. The increased expression of somatostatin receptors in IgA nephropathy kidney might be the potential pathogenesis of inflammatory renal disease. (J Histochem Cytochem 56:733–743, 2008)     

Somatostatin Analogues for Receptor Targeted Photodynamic Therapy

Photodynamic therapy (PDT) is an established treatment modality, used mainly for anticancer therapy that relies on the interaction of photosensitizer, light and oxygen. For the treatment of pathologies in certain anatomical sites, improved targeting of the photosensitizer is necessary to prevent damage to healthy tissue. We report on a novel dual approach of targeted PDT (vascular and cellular targeting) utilizing the expression of neuropeptide somatostatin receptor (sst₂) on tumor and neovascular-endothelial cells. We synthesized two conjugates containing the somatostatin analogue [Tyr3]-octreotate and Chlorin e6 (Ce6): Ce6-K₃-[Tyr3]-octreotate (1) and Ce6-[Tyr3]-octreotate-K₃-[Tyr3]-octreotate (2). Investigation of the uptake and photodynamic activity of conjugates in-vitro in human erythroleukemic K562 cells showed that conjugation of [Tyr3]-octreotate with Ce6 in conjugate 1 enhances uptake (by a factor 2) in cells over-expressing sst₂ compared to wild-type cells. Co-treatment with excess free Octreotide abrogated the phototoxicity of conjugate 1 indicative of a specific sst₂-mediated effect. In contrast conjugate 2 showed no receptor-mediated effect due to its high hydrophobicity. When compared with un-conjugated Ce6, the PDT activity of conjugate 1 was lower. However, it showed higher photostability which may compensate for its lower phototoxicity. Intra-vital fluorescence pharmacokinetic studies of conjugate 1 in rat skin-fold observation chambers transplanted with sst₂ ⁺ AR42J acinar pancreas tumors showed significantly different uptake profiles compared to free Ce6. Co-treatment with free Octreotide significantly reduced conjugate uptake in tumor tissue (by a factor 4) as well as in the chamber neo-vasculature. These results show that conjugate 1 might have potential as an in-vivo sst₂ targeting photosensitizer conjugate.     

Discovery of novel non-peptidic β-alanine piperazine amide derivatives and their optimization to achiral,easily accessible,potent and selective somatostatin sst1 receptor antagonists

Structural simplification of the core moieties of obeline and ergoline somatostatin sst₁ receptor antagonists, followed by systematic optimization, led to the identification of novel, highly potent and selective sst₁ receptor antagonists. These achiral, non-peptidic compounds are easily prepared and show promising PK properties in rodents.     

The Elusive Nature of Cerebellar Somatostatin Receptors: Studies in Rat,Monkey and Human Cerebellum

Abstract

The pharmacological profile and localization of somatostatin (SRIF) receptors were determined in rat, monkey and human cerebellum. In rat cerebellar cortex, low ss₁/sst₄, intermediate sst₂ and very high sst₃ receptor mRNA levels were found, sst₁ mRNA was also expressed in the deep cerebellar nuclei. [¹²⁵I]Tyr³-octreotide binding sites in cerebellar membranes correlated with recombinant sst₂, but not with sst₅ or sst₃ receptors and were found in the molecular layer of the cerebellum. [¹²⁵I]CGP 23996 (in Na⁺-buffer) binding in rat cerebellum correlated with sst₁ or sst₄, but not with sst₂, sst₃ or sst₅ receptor binding. Similar data were obtained in rhesus monkey cerebellum. mRNAs for all five receptors were found in the granule cell layer of the human cerebellum and/or in the dentate nucleus. [¹²⁵I]Tyr³-octreotide binding was strong in the molecular layer and correlated with that of recombinant sst₂ receptors, but not with sst₃ or sst₅ receptors. [¹²⁵I]CGP 23996 (in Mg⁺⁺-buffer) binding was heterogeneous (about 75%. to sst₂ and 25% to sst₁ and/or sst₄ receptors). The molecular and granular layers were equally and the dentate nucleus strongly labeled. Thus. SRIF receptors of the sst₂, sst₁ and/or sst₄ subtype are present in the rat, monkey and human cerebellum. In the latter two species, the sst₂ type appears to be predominant. Surprisingly, the high expression of sst₃ receptor mRNA is not supported by radioligand binding data in any of the species studied. The reason for this discrepancy remains to be elucidated.     

Expression of the Somatostatin Receptor Subtype 4 in Intact and Inflamed Pulmonary Tissues

Somatostatin released from capsaicin-sensitive sensory nerves of the lung during endotoxin-induced murine pneumonitis inhibits inflammation and hyperresponsiveness, presumably via somatostatin receptor subtype 4 (sst₄). The goal of the present study was to identify sst₄ receptors in mouse and human lungs and to reveal its inflammation-induced alterations with real-time quantitative PCR, Western blot, and immunohistochemistry. In non-inflamed mouse and human lungs, mRNA expression and immunolocalization of sst₄ are very similar. They are present on bronchial epithelial, vascular endothelial, and smooth-muscle cells. The sst₄ receptor protein in the mouse lung significantly increases 24 hr after intranasal endotoxin administration as well as in response to 3 months of whole-body cigarette smoke exposure, owing to the infiltrating sst₄-positivite mononuclear cells and neutrophils. In the chronically inflamed human lung, the large number of activated macrophages markedly elevate sst₄ mRNA levels, although there is no change in acute purulent pneumonia, in which granulocytes accumulate. Despite mouse granulocytes, human neutrophils do not show sst₄ immunopositivity. We provide the first evidence for the expression, localization, and inflammation-induced alterations of sst₄ receptors in murine and human lungs. Inasmuch as tissue distribution of this receptor is highly similar, extrapolation of murine experimental results to human conditions might be possible. (J Histochem Cytochem 57:1127–1137, 2009)     

Limitations of Current Approaches For The Treatment of Acromegaly

Objective: Acromegaly is a rare disease characterized by hypersecretion of growth hormone (GH), typically from a benign pituitary somatotroph adenoma, that leads to subsequent hypersecretion of insulin-like growth factor 1 (IGF-1). Patients with acromegaly have an increased risk of mortality and progressive worsening of comorbidities. Surgery, medical therapy, and radiotherapy are currently available treatment approaches for patients with acromegaly, with overall therapeutic goals of lowering GH levels and achieving normal IGF-1 levels, reducing tumor size, improving comorbidities, and minimizing mortality risk. Although surgery can lead to biochemical remission in some patients with acromegaly, many patients will continue to have uncontrolled disease and require additional treatment.Methods: We reviewed recently published reports and present a summary of the safety and efficacy of current treatment modalities for patients with acromegaly.Results: A substantial proportion of patients who receive medical therapy or radiotherapy will have persistently elevated GH and/or IGF-1. Because of the serious health consequences of continued elevation of GH and IGF-1, there is a need to improve therapeutic approaches to optimize biochemical control, particularly in high-need patient populations for whom current treatment options provide limited benefit.Conclusion: This review discusses current treatment options for patients with acromegaly, limitations associated with each treatment approach, and areas within the current treatment algorithm, as well as patient populations for which improved therapeutic options are needed. Novel agents in development were also highlighted, which have the potential to improve management of patients with uncontrolled or persistent acromegaly.Abbreviations:AACE = American Association of Clinical EndocrinologistsAE = adverse eventATG = AutogelCFRT = conventional fractionated radiotherapyDA = dopamine agonistENDO = Endocrine SocietyGH = growth hormoneGHRA = growth hormone receptor antagonistIGF-1 = insulin-like growth factor 1LAR = long-acting releaseLFT = liver function testSC = subcutaneousSRS = stereotactic radiosurgerySSA = somatostatin analoguesst = somatostatin receptorsst₂ = somatostatin receptor subtype 2sst₅ = somatostatin receptor subtype 5TSS = transsphenoidal surgery     

Functional effects of somatostatin receptor 1 activation on synaptic transmission in the mouse hippocampus

Somatostatin‐14 (SRIF) co‐localizes with GABA in the hippocampus and regulates neuronal excitability. A role of SRIF in the control of hippocampal activity has been proposed, although the exact contribution of each SRIF receptor (sst₁–sst₅) in mediating SRIF action requires some clarification. We used hippocampal slices of wild‐type and sst₁ knockout (KO) mice and selective pharmacological tools to provide conclusive evidence for a role of sst₁ in mediating SRIF inhibition of synaptic transmission. With single‐ and double‐label immunohistochemistry, we determined the distribution of sst₁ in hippocampal slices and we quantified sst₁ colocalization with SRIF. With electrophysiology, we found that sst₁ activation with CH‐275 inhibited both the NMDA‐ and the α‐amino‐3‐hydroxy‐5‐methyl‐4‐isoxazolepropionic acid (AMPA)‐mediated responses. Results from sst₁ KO slices confirmed the specificity of CH‐275 effects; sst₁ activation did not affect the inhibitory transmission which was in contrast increased by sst₄ activation with L‐803,087 in both wild‐type and sst₁ KO slices. The AMPA‐mediated responses were increased by L‐803,087. Functional interaction between sst₁ and sst₄ is suggested by the finding that their combined activation prevented the CH‐275‐induced inhibition of AMPA transmission. The involvement of pre‐synaptic mechanisms in mediating inhibitory effects of sst₁ on excitatory transmission was demonstrated by the finding that CH‐275 (i) increased the paired‐pulse facilitation ratio, (ii) did not influence the AMPA depolarization in the presence of tetrodotoxin, and (iii) inhibited glutamate release induced by epileptiform treatment. We conclude that SRIF control of excitatory transmission through an action at sst₁ may represent an important contribution to the regulation of hippocampal activity.     

Somatostatin receptor subtype 4 modulates L-type calcium channels via Gβγ and PKC signaling in rat retinal ganglion cells

Somatostatin subtype-4 receptors (sst₄) inhibit L-type calcium channel currents (I_Ca) in retinal ganglion cells (RGCs). Here we identify the signaling pathways involved in sst₄ stimulation leading to suppression of I_Ca in RGCs. Whole cell patch clamp recordings were made on isolated immunopanned RGCs using barium as a charge carrier to isolate I_Ca. Application of the selective sst₄ agonist, L-803 (10 nM), reduced I_Ca by 41.2%. Pretreatment of cells with pertussis toxin (Gi/o inhibitor) did not prevent the action of L-803, which reduced I_Ca by 34.7%. To determine the involvement of Gβγ subunits after sst₄ activation, depolarizing pre-pulse facilitation paradigms were used to remove voltage-dependent inhibition of calcium channels. Pre-pulse facilitation did not reverse the inhibitory effects of L-803 on I_Ca (8.4 vs. 8.8% reductions, ctrl vs. L-803); however, pharmacologic inhibition of Gβγ reduced I_Ca suppression by L-803 (23.0%, P < 0.05). Inhibition of PKC (GF109203X; GFX) showed a concentration-dependent effect in preventing the action of L-803 on I_Ca (1 μM GFX, 34.3%; 5 μM GFX, 14.6%, P < 0.05). When both PKC and Gβγ were inhibited, the effects of L-803 on I_Ca were blocked (1.8%, P < 0.05). These results suggest that sst₄ stimulation modulates RGC calcium channels via Gβγ and PKC activation. Since reducing intracellular Ca²⁺ is known to be neuroprotective in RGCs, modulating these sst₄ signaling pathways may provide insights to the discovery of unique therapeutic targets to reduce intracellular Ca²⁺ levels in RGCs.     

Tetraamine‐modified octreotide and octreotate: labeling with 99mTc and preclinical comparison in AR4‐2J cells and AR4‐2J tumor‐bearing mice

Two somatostatin analogues, [^99mTc]Demotide and [^99mTc]Demotate 4, were compared with [^99mTc]Demotate 1, a previously reported somatostatin receptor subtype 2 (sst₂) targeting tracer. Conjugates were prepared by coupling an open‐chain tetraamine chelator to D ‐Phe¹ of [Tyr³]‐octreotide or [Tyr³]‐octreotate, respectively, via a p‐benzylaminodiglycolic acid spacer adopting solid‐phase peptide synthesis techniques. Peptide conjugates were collected in a highly pure form after chromatographic purification. Eventually, [^99mTc]Demotide and [^99mTc]Demotate 4 were obtained in ～1 Ci/µmol specific activity and >96% purity after labeling under alkaline conditions. Demotide and Demotate 4 exhibited similar high binding affinities for the sst₂ expressed in AR4‐2J cells with IC₅₀ values 0.16 and 0.10 nM, respectively. The (radio)metallated analogues [^99mTc]Demotide and [^99mTc]Demotate 4 showed equally high affinities to the sst₂ during saturation binding assays in AR4‐2J cell membranes (K_ds 0.08 and 0.07 nM, respectively). During incubation at 37 °C with AR4‐2J cells, the radiopeptides internalized effectively via a receptor‐mediated process, with [^99mTc]Demotate 4 exhibiting a faster internalization rate than [^99mTc]Demotide. After injection in athymic mice bearing sst₂‐expressing AR4‐2J tumors, the radiotracers showed high and specific uptake in the tumor (>25%ID/g at 1 h) and in the sst₂–positive organs. However, both [^99mTc]Demotide and [^99mTc]Demotate 4 showed unfavorably higher background activity, especially in the abdomen, in comparison to [^99mTc]Demotate 1 and are, therefore, less suited than [^99mTc]Demotate 1 for sst₂‐targeted tumor imaging in man. Copyright © 2005 European Peptide Society and John Wiley & Sons, Ltd.     

Somatostatin receptors

In 1972, Brazeau et al. isolated somatostatin (somatotropin release-inhibiting factor, SRIF), a cyclic polypeptide with two biologically active isoforms (SRIF-14 and SRIF-28). This event prompted the successful quest for SRIF receptors. Then, nearly a quarter of a century later, it was announced that a neuropeptide, to be named cortistatin (CST), had been cloned, bearing strong resemblance to SRIF. Evidence of special CST receptors never emerged, however. CST rather competed with both SRIF isoforms for specific receptor binding. And binding to the known subtypes with affinities in the nanomolar range, it has therefore been acknowledged to be a third endogenous ligand at SRIF receptors.This review goes through mechanisms of signal transduction, pharmacology, and anatomical distribution of SRIF receptors. Structurally, SRIF receptors belong to the superfamily of G protein-coupled (GPC) receptors, sharing the characteristic seven-transmembrane-segment (STMS) topography. Years of intensive research have resulted in cloning of five receptor subtypes (sst₁-sst₅), one of which is represented by two splice variants (sst_2A and sst_2B). The individual subtypes, functionally coupled to the effectors of signal transduction, are differentially expressed throughout the mammalian organism, with corresponding differences in physiological impact. It is evident that receptor function, from a physiological point of view, cannot simply be reduced to the accumulated operations of individual receptors. Far from being isolated functional units, receptors co-operate. The total receptor apparatus of individual cell types is composed of different-ligand receptors (e.g. SRIF and non-SRIF receptors) and co-expressed receptor subtypes (e.g. sst₂ and sst₅ receptors) in characteristic proportions. In other words, levels of individual receptor subtypes are highly cell-specific and vary with the co-expression of different-ligand receptors. However, the question is how to quantify the relative contributions of individual receptor subtypes to the integration of transduced signals, ultimately the result of collective receptor activity. The generation of knock-out (KO) mice, intended as a means to define the contributions made by individual receptor subtypes, necessarily marks but an approximation. Furthermore, we must now take into account the stunning complexity of receptor co-operation indicated by the observation of receptor homo- and heterodimerisation, let alone oligomerisation. Theoretically, this phenomenon adds a novel series of functional megareceptors/super-receptors, with varied pharmacological profiles, to the catalogue of monomeric receptor subtypes isolated and cloned in the past. SRIF analogues include both peptides and non-peptides, receptor agonists and antagonists. Relatively long half lives, as compared to those of the endogenous ligands, have been paramount from the outset. Motivated by theoretical puzzles or the shortcomings of present-day diagnostics and therapy, investigators have also aimed to produce subtype-selective analogues. Several have become available.     

Characterization of binding kinetics of A2AR to Gαs protein by surface plasmon resonance

Because of their surface localization, G protein-coupled receptors (GPCRs) are often pharmaceutical targets as they respond to a variety of extracellular stimuli (e.g., light, hormones, small molecules) that may activate or inhibit a downstream signaling response. The adenosine A_2A receptor (A_2AR) is a well-characterized GPCR that is expressed widely throughout the human body, with over 10 crystal structures determined. Truncation of the A_2AR C-terminus is necessary for crystallization as this portion of the receptor is long and unstructured; however, previous work suggests shortening of the A_2AR C-terminus from 412 to 316 amino acids (A_2AΔ316R) ablates downstream signaling, as measured by cAMP production, to below that of constitutive full-length A_2AR levels. As cAMP production is downstream of the first activation event—coupling of G protein to its receptor—investigating that first step in activation is important in understanding how the truncation effects native GPCR function. Here, using purified receptor and Gα_s proteins, we characterize the association of A_2AR and A_2AΔ316R to Gα_s with and without GDP or GTPγs using surface plasmon resonance (SPR). Gα_s affinity for A_2AR was greatest for apo-Gα_s, moderately affected in the presence of GDP and nearly completely ablated by the addition of GTPγs. Truncation of the A_2AR C-terminus (A_2AΔ316R) decreased the affinity of the unliganded receptor for Gα_s by ～20%, suggesting small changes to binding can greatly impact downstream signaling.     

Advances in understanding neuronal somatostatin receptors

It has long been considered that somatostatin acts as a neuromodulator in the mammalian central nervous system but its precise physiological roles remain elusive. Early studies to identify somatostatin-binding sites revealed a widespread heterogeneous pattern, especially in the CNS. More recently, a family of somatostatin receptors have been identified, of which five genes (sst_1–5) have been cloned. In this review, we discuss current data describing the localisation of the five receptor types. Recent progress in understanding their function has been made using high-affinity, selective receptor ligands and transgenic animal technology. Finally, the therapeutic potential for somatostatin receptor-selective compounds as analgesics is considered.     

Expression of Activin Receptor-like Kinase 7 in Adipose Tissues

The tissue distribution of activin receptor-like kinase 7 (Alk7) expression, the signaling ability of Alk7 variants, and Alk7 expression in response to β₃-adrenergic receptor activation were examined. Expression levels of Alk7 varied greatly among tissues but were highest in white adipose tissue and brown adipose tissue. In addition to full-length Alk7 (Alk7-v1), Alk7-v3, an Alk7 variant, was expressed in adipose tissues, brain, and ovary. Nodal transmits signals via Alk7 in cooperation with its coreceptor, Cripto. Evaluation of the ability of Alk7 variants to confer Nodal signaling using luciferase-based reporter assays showed that Alk7-v3 does not transmit Nodal-Cripto-mediated signals. Expression of Alk7 was down-regulated in brown but not in white adipose tissue treated with CL316,243, a β₃-adrenergic receptor agonist. These results suggest involvement of Alk7 in modulation of metabolism in the adipose tissues in response to β₃-adrenergic receptor activation.     

Somatostatin receptor subtype 4 modulates L-type calcium channels via Gβγ and PKC signaling in rat retinal ganglion cells

Somatostatin subtype-4 receptors (sst₄) inhibit L-type calcium channel currents (I_Ca) in retinal ganglion cells (RGCs). Here we identify the signaling pathways involved in sst₄ stimulation leading to suppression of I_Ca in RGCs. Whole cell patch clamp recordings were made on isolated immunopanned RGCs using barium as a charge carrier to isolate I_Ca. Application of the selective sst₄ agonist, L-803 (10 nM), reduced I_Ca by 41.2%. Pretreatment of cells with pertussis toxin (Gi/o inhibitor) did not prevent the action of L-803, which reduced I_Ca by 34.7%. To determine the involvement of Gβγ subunits after sst₄ activation, depolarizing pre-pulse facilitation paradigms were used to remove voltage-dependent inhibition of calcium channels. Pre-pulse facilitation did not reverse the inhibitory effects of L-803 on I_Ca (8.4 vs. 8.8% reductions, ctrl vs. L-803); however, pharmacologic inhibition of Gβγ reduced I_Ca suppression by L-803 (23.0%, P < 0.05). Inhibition of PKC (GF109203X; GFX) showed a concentration-dependent effect in preventing the action of L-803 on I_Ca (1 μM GFX, 34.3%; 5 μM GFX, 14.6%, P < 0.05). When both PKC and Gβγ were inhibited, the effects of L-803 on I_Ca were blocked (1.8%, P < 0.05). These results suggest that sst₄ stimulation modulates RGC calcium channels via Gβγ and PKC activation. Since reducing intracellular Ca²⁺ is known to be neuroprotective in RGCs, modulating these sst₄ signaling pathways may provide insights to the discovery of unique therapeutic targets to reduce intracellular Ca²⁺ levels in RGCs.     

Somatostatin increases mitogen-induced IL-2 secretion and proliferation of human jurkat T cells via sst3 receptor isotype

The neuropeptide somatostatin (SRIF) modulates normal and leukemia T cell proliferation. However, neither molecular isotypes of receptors nor mechanisms involved in these somatostatin actions have been elucidated as yet. Here we show by using RT-PCR approach that mitogen-activated leukemia T cells (Jurkat) express mRNA for a single somatostatin receptor, sst3. This mRNA is apparently translated into protein since specific somatostatin binding sites (K_I1 = 78 ± 3 pM) were detected in semipurified plasma membrane preparations by using ¹²⁵I-Tyr¹-SRIF14 as a radioligand. Moreover, somatostatin inhibits adenylyl cyclase activity with similar efficiency (IC₅₀ = 23 ± 4 pM) thus strongly suggesting a functional coupling of sst3 receptor to this transduction pathway. The involvement of sst3 receptor in immuno-modulatory actions of somatostatin was assessed by analysis of neuropeptide effects on IL-2 secretion and on proliferation of mitogen-activated Jurkat cells. Our data show that in the concentrations comprised between 10 pM and 10 nM, somatostatin potentiates IL-2 secretion. This effect is correlated with somatostatin-dependent increase of Jurkat cell proliferation since the EC₅₀ concentrations for both actions were almost identical (EC₅₀ = 22 ± 9 pM and EC₅₀ = 12 ± 1 pM for IL-2 secretion and proliferation, respectively). Altogether, these data strongly suggest that in mitogen-activated Jurkat cells, somatostatin increases cell proliferation through the increase of IL-2 secretion via a functional sst3 receptor negatively coupled to the adenylyl cyclase pathway. J. Cell. Biochem. 68:62–73, 1998. © 1998 Wiley-Liss, Inc.     

Pannexin 1 Channels Link Chemoattractant Receptor Signaling to Local Excitation and Global Inhibition Responses at the Front and Back of Polarized Neutrophils

Neutrophil chemotaxis requires excitatory signals at the front and inhibitory signals at the back of cells, which regulate cell migration in a chemotactic gradient field. We have previously shown that ATP release via pannexin 1 (PANX1) channels and autocrine stimulation of P2Y₂ receptors contribute to the excitatory signals at the front. Here we show that PANX1 also contributes to the inhibitory signals at the back, namely by providing the ligand for A_2A adenosine receptors. In resting neutrophils, we found that A_2A receptors are uniformly distributed across the cell surface. In polarized cells, A_2A receptors redistributed to the back where their stimulation triggered intracellular cAMP accumulation and protein kinase A (PKA) activation, which blocked chemoattractant receptor signaling. Inhibition of PANX1 blocked A_2A receptor stimulation and cAMP accumulation in response to formyl peptide receptor stimulation. Treatments that blocked endogenous A_2A receptor signaling impaired the polarization and migration of neutrophils in a chemotactic gradient field and resulted in enhanced ERK and p38 MAPK signaling in response to formyl peptide receptor stimulation. These findings suggest that chemoattractant receptors require PANX1 to trigger excitatory and inhibitory signals that synergize to fine-tune chemotactic responses at the front and back of neutrophils. PANX1 channels thus link local excitatory signals to the global inhibitory signals that orchestrate chemotaxis of neutrophils in gradient fields.