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ß-Adrenergic receptor signaling increases NAADP and cADPR levels in the heart
Evidence suggests that ß-Adrenergic receptor signaling increases heart rate and force through not just cyclic AMP but also the Ca 2+-releasing second messengers NAADP (nicotinic acid adenine dinucleotide phosphate) and cADPR (cyclic ADP-ribose). Nevertheless, proof of the physiological relevance of these messengers requires direct measurements of their levels in response to receptor stimulation. Here we report that in intact Langendorff-perfused hearts ß-adrenergic stimulation increased both messengers, with NAADP being transient and cADPR being sustained. Both NAADP and cADPR have physiological and therefore pathological relevance by providing alternative drug targets in the ß-adrenergic receptor signaling pathway. © 2012 Elsevier Inc.
β-Adrenergic receptor signaling increases NAADP and cADPR levels in the heart.
Evidence suggests that β-Adrenergic receptor signaling increases heart rate and force through not just cyclic AMP but also the Ca(2+)-releasing second messengers NAADP (nicotinic acid adenine dinucleotide phosphate) and cADPR (cyclic ADP-ribose). Nevertheless, proof of the physiological relevance of these messengers requires direct measurements of their levels in response to receptor stimulation. Here we report that in intact Langendorff-perfused hearts β-adrenergic stimulation increased both messengers, with NAADP being transient and cADPR being sustained. Both NAADP and cADPR have physiological and therefore pathological relevance by providing alternative drug targets in the β-adrenergic receptor signaling pathway.
NAADP activates two-pore channels on T cell cytolytic granules to stimulate exocytosis and killing.
A cytotoxic T lymphocyte (CTL) kills an infected or tumorigenic cell by Ca(2+)-dependent exocytosis of cytolytic granules at the immunological synapse formed between the two cells. Although inositol 1,4,5-trisphosphate (IP(3))-mediated Ca(2+) release from the endoplasmic reticulum activates the store-operated Ca(2+)-influx pathway that is necessary for exocytosis, it is not a sufficient stimulus. Here we identify the Ca(2+)-mobilizing messenger nicotinic acid adenine dinucleotide phosphate (NAADP) and its recently identified molecular target, two-pore channels (TPCs), as being important for T cell receptor signaling in CTLs. We demonstrate that cytolytic granules are not only reservoirs of cytolytic proteins but are also the acidic Ca(2+) stores mobilized by NAADP via TPC channels on the granules themselves, so that TPCs migrate to the immunological synapse upon CTL activation. Moreover, NAADP activates TPCs to drive exocytosis in a way that is not mimicked by global Ca(2+) signals induced by IP(3) or ionomycin, suggesting that critical, local Ca(2+) nanodomains around TPCs stimulate granule exocytosis. Hence, by virtue of the NAADP/TPC pathway, cytolytic granules generate Ca(2+) signals that lead to their own exocytosis and to cell killing. This study highlights a selective role for NAADP in stimulating exocytosis crucial for immune cell function and may impact on stimulus-secretion coupling in wider cellular contexts.
Phospholipase C-dependent Ca2+ release by worm and mammal sperm factors.
Egg activation in all animals evidently requires the synthesis of inositol 1,4,5-trisphosphate (InsP(3)) from phosphatidylinositol 4,5-bisphosphate (PIP(2)) by phospholipase C (PLC). Depending on the organism, InsP(3) elicits either calcium oscillations or a single wave, which in turn initiates development. A soluble component in boar sperm that activates mammalian eggs has been suggested to be a PLC isoform. We tested this hypothesis in vitro using egg microsomes of Chaetopterus. Boar sperm factor elicited Ca(2+) release from the microsomes by an InsP(3)-dependent mechanism. The PLC inhibitor U-73122, but not its inactive analog U-73343, blocked the response to sperm factor but not to InsP(3). U-73122 also inhibited the activation of fertilized and parthenogenetic eggs. Chaetopterus sperm also contained a similar activity. These results strongly support the hypothesis that sperm PLCs are ubiquitous mediators of egg activation at fertilization.
Calcium release from the endoplasmic reticulum of higher plants elicited by the NADP metabolite nicotinic acid adenine dinucleotide phosphate.
Higher plants share with animals a responsiveness to the Ca(2+) mobilizing agents inositol 1,4,5-trisphosphate (InsP(3)) and cyclic ADP-ribose (cADPR). In this study, by using a vesicular (45)Ca(2+) flux assay, we demonstrate that microsomal vesicles from red beet and cauliflower also respond to nicotinic acid adenine dinucleotide phosphate (NAADP), a Ca(2+)-releasing molecule recently described in marine invertebrates. NAADP potently mobilizes Ca(2+) with a K(1/2) = 96 nM from microsomes of nonvacuolar origin in red beet. Analysis of sucrose gradient-separated cauliflower microsomes revealed that the NAADP-sensitive Ca(2+) pool was derived from the endoplasmic reticulum. This exclusively nonvacuolar location of the NAADP-sensitive Ca(2+) pathway distinguishes it from the InsP(3)- and cADPR-gated pathways. Desensitization experiments revealed that homogenates derived from cauliflower tissue contained low levels of NAADP (125 pmol/mg) and were competent in NAADP synthesis when provided with the substrates NADP and nicotinic acid. NAADP-induced Ca(2+) release is insensitive to heparin and 8-NH(2)-cADPR, specific inhibitors of the InsP(3)- and cADPR-controlled mechanisms, respectively. However, NAADP-induced Ca(2+) release could be blocked by pretreatment with a subthreshold dose of NAADP, as previously observed in sea urchin eggs. Furthermore, the NAADP-gated Ca(2+) release pathway is independent of cytosolic free Ca(2+) and therefore incapable of operating Ca(2+)-induced Ca(2+) release. In contrast to the sea urchin system, the NAADP-gated Ca(2+) release pathway in plants is not blocked by L-type channel antagonists. The existence of multiple Ca(2+) mobilization pathways and Ca(2+) release sites might contribute to the generation of stimulus-specific Ca(2+) signals in plant cells.
Cyclic ADP ribose as a calcium-mobilizing messenger.
This Perspective by Galione and Churchill is one in a series on intracellular calcium release mechanisms. The authors review the evidence for cyclic adenosine diphosphate ribose (cADPR) being a second messenger involved in regulating intracellular calcium. In addition, the physiological stimuli and responses mediated by cADPR are discussed. The Perspective is accompanied by a movie showing a calcium wave triggered by cADPR.
NAADP controls cross-talk between distinct Ca2+ stores in the heart.
In cardiac muscle the sarcoplasmic reticulum (SR) plays a key role in the control of contraction, releasing Ca(2+) in response to Ca(2+) influx across the sarcolemma via voltage-gated Ca(2+) channels. Here we report evidence for an additional distinct Ca(2+) store and for actions of nicotinic acid adenine dinucleotide phosphate (NAADP) to mobilize Ca(2+) from this store, leading in turn to enhanced Ca(2+) loading of the SR. Photoreleased NAADP increased Ca(2+) transients accompanying stimulated action potentials in ventricular myocytes. The effects were prevented by bafilomycin A (an H(+)-ATPase inhibitor acting on acidic Ca(2+) stores), by desensitizing concentrations of NAADP, and by ryanodine and thapsigargin to suppress SR function. Bafilomycin A also suppressed staining of acidic stores with Lysotracker Red without affecting SR integrity. Cytosolic application of NAADP by means of its membrane permeant acetoxymethyl ester increased myocyte contraction and the frequency and amplitude of Ca(2+) sparks, and these effects were inhibited by bafilomycin A. Effects of NAADP were associated with an increase in SR Ca(2+) load and appeared to be regulated by beta-adrenoreceptor stimulation. The observations are consistent with a novel role for NAADP in cardiac muscle mediated by Ca(2+) release from bafilomycin-sensitive acidic stores, which in turn enhances SR Ca(2+) release by increasing SR Ca(2+) load.
TPCs: Endolysosomal channels for Ca2+ mobilization from acidic organelles triggered by NAADP.
Two-pore channels (TPCs or TPCNs) are novel members of the large superfamily of voltage-gated cation channels with slightly higher sequence homology to the pore-forming subunits of voltage-gated Ca(2+) and Na(+) channels than most other members. Recent studies demonstrate that TPCs locate to endosomes and lysosomes and form Ca(2+) release channels that respond to activation by the Ca(2+) mobilizing messenger, nicotinic acid adenine dinucleotide phosphate (NAADP). With multiple endolysosomal targeted NAADP receptors now identified, important new insights into the regulation of endolysosomal function in health and disease will therefore be unveiled.
Nicotinic acid adenine dinucleotide phosphate regulates skeletal muscle differentiation via action at two-pore channels.
Calcium signaling is essential for the differentiation of many cell types, including skeletal muscle cells, but its mechanisms remain elusive. Here we demonstrate a crucial role for nicotinic acid adenine dinucleotide phosphate (NAADP) signaling in skeletal muscle differentiation. Although the inositol trisphosphate pathway may have a partial role to play in this process, the ryanodine signaling cascade is not involved. In both skeletal muscle precursors and C2C12, cells interfering with NAADP signaling prevented differentiation, whereas promoting NAADP signaling potentiated differentiation. Moreover, siRNA knockdown of two-pore channels, the target of NAADP, attenuated differentiation. The data presented here strongly suggest that in myoblasts, NAADP acts at acidic organelles on the recently discovered two-pore channels to promote differentiation.
Unique kinetics of nicotinic acid-adenine dinucleotide phosphate (NAADP) binding enhance the sensitivity of NAADP receptors for their ligand.
Nicotinic acid-adenine dinucleotide phosphate (NAADP) is a novel and potent Ca(2+)-mobilizing agent in sea urchin eggs and other cell types. Little is known, however, concerning the properties of the putative intracellular NAADP receptor. In the present study we have characterized NAADP binding sites in sea urchin egg homogenates. [(32)P]NAADP bound to a single class of high-affinity sites that were reversibly inhibited by NaCl but insensitive to pH and Ca(2+). Binding of [(32)P]NAADP was lost in preparations that did not mobilize Ca(2+) in response to NAADP, indicating that [(32)P]NAADP probably binds to a receptor mediating Ca(2+) mobilization. Addition of excess unlabelled NAADP, at various times after initiation of [(32)P]NAADP binding, did not result in displacement of bound [(32)P]NAADP. These data show that NAADP becomes irreversibly bound to its receptor immediately upon association. Accordingly, incubation of homogenates with low concentrations of NAADP resulted in maximal labelling of NAADP binding sites. This unique property renders NAADP receptors exquisitely sensitive to their ligand, thereby allowing detection of minute changes in NAADP levels.
Solubilization of receptors for the novel Ca2+-mobilizing messenger, nicotinic acid adenine dinucleotide phosphate.
Nicotinic acid adenine dinucleotide phosphate (NAADP) is a potent Ca(2+) mobilizing agent in a variety of broken and intact cell preparations. In sea urchin egg homogenates, NAADP releases Ca(2+) independently of inositol trisphosphate or ryanodine receptor activation. Little, however, is known concerning the molecular target for NAADP. Here we report for the first time solubilization of NAADP receptors from sea urchin egg homogenates. Supernatant fractions, prepared following Triton X-100 treatment, bound [(32)P]NAADP with similar affinity and selectivity as membrane preparations. Furthermore, the unusual non-dissociating nature of NAADP binding to its receptor was preserved upon solubilization. NAADP receptors could also be released into supernatant fractions upon detergent treatment of membranes prelabeled with [(32)P]NAADP. Tagged receptors prepared in this way, were readily resolved by native gel electrophoresis as a single protein target. Gel filtration and sucrose density gradient centrifugation analysis indicates that NAADP receptors are substantially smaller than inositol trisphosphate or ryanodine receptors, providing further biochemical evidence that NAADP activates a novel intracellular Ca(2+) release channel.
Pharmacological characterization of the putative cADP-ribose receptor.
cADP-ribose (cADPR), a naturally occurring metabolite of NAD(+), has been shown to be an important regulator of intracellular Ca(2+) release. Considerable evidence suggests that cADPR is the endogenous modulator of the ryanodine receptor (RyR), which mediates Ca(2+)-induced Ca(2+) release (CICR). Indeed, cADPR-mediated Ca(2+) release is subject to functional regulation by other modulators of CICR, including Ca(2+), caffeine and calmodulin. However, the underlying basis behind the effect of such agents on cADPR activity (in particular whether they regulate cADPR binding), as well as the precise nature of the cADPR receptor remains unclear. In the present study, use of (32)P-radiolabelled cADPR has enabled a detailed pharmacological characterization of cADPR-binding sites in sea urchin egg homogenates. We report that cADPR binds specifically to a single class of high affinity receptor. Retainment of binding to membranes after a high-salt wash suggests the involvement of either an integral membrane protein (possibly the RyR itself) or a peripheral protein tightly associated to the membrane. Insensitivity of [(32)P]cADPR binding to either FK506 or rapamycin suggests that this does not concern the FK506-binding protein. Significantly, binding is highly robust, being relatively insensitive to both endogenous and pharmacological modulators of RyR-mediated CICR. In turn, this suggests that such agents modulate cADPR-mediated Ca(2+) release primarily by tuning the 'gain' of the CICR system, upon which cADPR acts, rather than influencing the interaction of cADPR with its target receptor. The exception to this is calmodulin, for which our results indicate an additional role in facilitating cADPR binding.
Ca2+ release from the endoplasmic reticulum of NY-ESO-1-specific T cells is modulated by the affinity of TCR and by the use of the CD8 coreceptor.
Although several cancer immunotherapy strategies are based on the use of analog peptides and on the modulation of the TCR affinity of adoptively transferred T cells, it remains unclear whether tumor-specific T cell activation by strong and weak TCR stimuli evoke different Ca(2+) signatures from the Ca(2+) intracellular stores and whether the amplitude of Ca(2+) release from the endoplasmic reticulum (ER) can be further modulated by coreceptor binding to peptide/MHC. In this study, we combined functional, structural, and kinetic measurements to correlate the intensity of Ca(2+) signals triggered by the stimulation of the 1G4 T cell clone specific to the tumor epitope NY-ESO-1(157-165). Two analogs of the NY-ESO-1(157-165) peptide, having similar affinity to HLA-A2 molecules, but a 6-fold difference in binding affinity for the 1G4 TCR, resulted in different Ca(2+) signals and T cell activation. 1G4 stimulation by the stronger stimulus emptied the ER of stored Ca(2+), even in the absence of CD8 binding, resulting in sustained Ca(2+) influx. In contrast, the weaker stimulus induced only partial emptying of stored Ca(2+), resulting in significantly diminished and oscillatory Ca(2+) signals, which were enhanced by CD8 binding. Our data define the range of TCR/peptide MHC affinities required to induce depletion of Ca(2+) from intracellular stores and provide insights into the ability of T cells to tailor the use of the CD8 coreceptor to enhance Ca(2+) release from the ER. This, in turn, modulates Ca(2+) influx from the extracellular environment, ultimately controlling T cell activation.
Aplysia californica mediated cyclisation of novel 3'-modified NAD+ analogues: a role for hydrogen bonding in the recognition of cyclic adenosine 5'-diphosphate ribose.
Cyclic ADP-ribose mobilizes intracellular Ca2+ in a variety of cells. To elucidate the nature of the interaction between the C3' substituent of cADP-ribose and the cADPR receptor, three analogues of NAD+ modified in the adenosine ribase (xyloNAD+ 3'F-xyloNAD+ and 3'F-NAD+ were chemically synthesised from D-xylose and adenine starting materials. 3'F-NAD+ was readily converted to cyclic 3'F-ADP ribose by the action of the cyclase enzyme derived from the mollusc Aplysia californica. XyloNAD+ and 3'F-xyloNAD+ were cyclised only reluctantly and in poor yield to afford unstable cyclic products. Biological evaluation of cyclic 3'F-ADP ribose for calcium release in sea urchin egg homogenate gave an EC(50) of 1.5+/-0.5 microM. This high value suggests that the ability of the C3' substituent to donate a hydrogen bond is crucial for agonism.
Vasodilation by the calcium-mobilizing messenger cyclic ADP-ribose.
In artery smooth muscle, adenylyl cyclase-coupled receptors such as beta-adrenoceptors evoke Ca(2+) signals, which open Ca(2+)-activated potassium (BK(Ca)) channels in the plasma membrane. Thus, blood pressure may be lowered, in part, through vasodilation due to membrane hyperpolarization. The Ca(2+) signal is evoked via ryanodine receptors (RyRs) in sarcoplasmic reticulum proximal to the plasma membrane. We show here that cyclic adenosine diphosphate-ribose (cADPR), by activating RyRs, mediates, in part, hyperpolarization and vasodilation by beta-adrenoceptors. Thus, intracellular dialysis of cADPR increased the cytoplasmic Ca(2+) concentration proximal to the plasma membrane in isolated arterial smooth muscle cells and induced a concomitant membrane hyperpolarization. Smooth muscle hyperpolarization mediated by cADPR, by beta-adrenoceptors, and by cAMP, respectively, was abolished by chelating intracellular Ca(2+) and by blocking RyRs, cADPR, and BK(Ca) channels with ryanodine, 8-amino-cADPR, and iberiotoxin, respectively. The cAMP-dependent protein kinase A antagonist N-(2-[p-bromocinnamylamino]ethyl)-5-isoquinolinesulfonamide hydrochloride (H89) blocked hyperpolarization by isoprenaline and cAMP, respectively, but not hyperpolarization by cADPR. Thus, cADPR acts as a downstream element in this signaling cascade. Importantly, antagonists of cADPR and BK(Ca) channels, respectively, inhibited beta-adrenoreceptor-induced artery dilation. We conclude, therefore, that relaxation of arterial smooth muscle by adenylyl cyclase-coupled receptors results, in part, from a cAMP-dependent and protein kinase A-dependent increase in cADPR synthesis, and subsequent activation of sarcoplasmic reticulum Ca(2+) release via RyRs, which leads to activation of BK(Ca) channels and membrane hyperpolarization.
Nicotinic acid adenine dinucleotide phosphate mediates Ca2+ signals and contraction in arterial smooth muscle via a two-pool mechanism.
Previous studies of arterial smooth muscle have shown that inositol 1,4,5-trisphosphate (IP3) and cyclic ADP-ribose mobilize Ca2+ from the sarcoplasmic reticulum. In contrast, little is known about Ca2+ mobilization by nicotinic acid adenine dinucleotide phosphate, a pyridine nucleotide derived from beta-NADP+. We show here that intracellular dialysis of nicotinic acid adenine dinucleotide phosphate (NAADP) induces spatially restricted "bursts" of Ca2+ release that initiate a global Ca2+ wave and contraction in pulmonary artery smooth muscle cells. Depletion of sarcoplasmic reticulum Ca2+ stores with thapsigargin and inhibition of ryanodine receptors with ryanodine, respectively, block the global Ca2+ waves by NAADP. Under these conditions, however, localized Ca2+ bursts are still observed. In contrast, xestospongin C, an IP3 receptor antagonist, had no effect on Ca2+ signals by NAADP. We propose that NAADP mobilizes Ca2+ via a 2-pool mechanism, and that initial Ca2+ bursts are amplified by subsequent sarcoplasmic reticulum Ca2+ release via ryanodine receptors but not via IP3 receptors.
Analogues of the nicotinic acid adenine dinucleotide phosphate (NAADP) antagonist Ned-19 indicate two binding sites on the NAADP receptor.
Nicotinic acid adenine dinucleotide phosphate (NAADP) is a Ca(2+)-releasing messenger. Biological data suggest that its receptor has two binding sites: one high-affinity locking site and one low-affinity opening site. To directly address the presence and function of these putative binding sites, we synthesized and tested analogues of the NAADP antagonist Ned-19. Ned-19 itself inhibits both NAADP-mediated Ca(2+) release and NAADP binding. A fluorometry bioassay was used to assess NAADP-mediated Ca(2+) release, whereas a radioreceptor assay was used to assess binding to the NAADP receptor (only at the high-affinity site). In Ned-20, the fluorine is para rather than ortho as in Ned-19. Ned-20 does not inhibit NAADP-mediated Ca(2+) release but inhibits NAADP binding. Conversely, Ned-19.4 (a methyl ester of Ned-19) inhibits NAADP-mediated Ca(2+) release but cannot inhibit NAADP binding. Furthermore, Ned-20 prevents the self-desensitization response characteristic of NAADP in sea urchin eggs, confirming that this response is mediated by a high-affinity allosteric site to which NAADP binds in the radioreceptor assay. Collectively, these data provide the first direct evidence for two binding sites (one high- and one low-affinity) on the NAADP receptor.
Diabetes mellitus as a predictor for radial artery vasoreactivity in patients undergoing coronary artery bypass grafting.
OBJECTIVES: Our purpose was to examine the impact of diabetes mellitus (DM) on vasoreactivity and endothelial function of radial artery (RA) grafts ex vivo. BACKGROUND: The arteriopathy associated with DM may influence the surgeon's choice of conduits for revascularization. Arterial conduits and especially the RA are prone to vasospasm in the perioperative period. METHODS: The study population consisted of 98 patients with coronary artery disease undergoing coronary artery bypass grafting by using RA grafts. The maximum contractions of RA segments induced by K+ (66 mmol/l) and clinically important vasoconstrictors such as adrenaline (5 x 10(-5) mol/l), angiotensin II (10(-6) mol/l), and prostaglandin F2alpha (PGF2alpha) (10(-6) mol/l) were recorded. Relaxation of RA rings to carbachol (10(-4) mol/l) was used as a measure of endothelial function. Multivariate analysis was then applied to determine the role of clinical characteristics on the vasomotor responses to these agents. RESULTS: Vessels from patients with DM had greater contractions in response to adrenaline (p < 0.05), angiotensin (p < 0.05), and PGF2alpha (p < 0.01) compared with non-DM vessels, despite the similar vasoconstrictions induced by high K+ (p = NS). Diabetes mellitus was also associated with smaller vasorelaxations in response to carbachol (p < 0.001). In multivariate analysis, DM was an independent predictor of RA contractions in response to adrenaline (beta [SE] 3.085 [1.410], p = 0.031), angiotensin II (beta [SE] 3.838 [1.552], p = 0.015), and PGF2alpha (beta [SE] 4.609 [1.908], p = 0.018) but not K+ (p = NS). Diabetes mellitus was also independently associated with the vasorelaxations in response to carbachol (beta [SE] -15.645 [2.622], p = 0.0001). CONCLUSIONS: Diabetes mellitus is associated with impaired endothelial function and greater contractions of RA grafts in response to all of the clinically relevant vasoconstrictors. These findings suggest that the RA of diabetic patients may be more prone to spasm in response to endogenous vasoconstrictors, an observation with important implications for surgeons' choice of conduits in this cohort of patients.
The calcium-mobilizing messenger nicotinic acid adenine dinucleotide phosphate participates in sperm activation by mediating the acrosome reaction.
Before a sperm can fertilize an egg it must undergo a final activation step induced by the egg termed the acrosome reaction. During the acrosome reaction a lysosome-related organelle, the acrosome, fuses with the plasma membrane to release hydrolytic enzymes and expose an egg-binding protein. Because NAADP (nicotinic acid adenine dinucleotide phosphate) releases Ca(2+) from acidic lysosome-related organelles in other cell types, we investigated a possible role for NAADP in mediating the acrosome reaction. We report that NAADP binds with high affinity to permeabilized sea urchin sperm. Moreover, we used Mn(2+) quenching of luminal fura-2 and (45)Ca(2+) to directly demonstrate NAADP regulation of a cation channel on the acrosome. Additionally, we show that NAADP synthesis occurs through base exchange and is driven by an increase in Ca(2+). We propose a new model for acrosome reaction signaling in which Ca(2+) influx initiated by egg jelly stimulates NAADP synthesis and that this NAADP acts on its receptor/channel on the acrosome to release Ca(2+) to drive acrosomal exocytosis.
Calmodulin dissociation mediates desensitization of the cADPR-induced Ca2+ release mechanism.
Ryanodine receptor (RyR) activation by cyclic ADP-ribose (cADPR) is followed by homologous desensitization. Though poorly understood, this "switching off" process has provided a key experimental tool for determining the pathway through which cADPR mediates Ca(2+) release. Moreover, desensitization is likely to play an important role in shaping the complexities of Ca(2+) signaling involving cADPR, for example, localized release events and propagated waves. Using the sea urchin egg, we unmask a role of calmodulin, a component of the RyR complex and a key cofactor for cADPR activity, during RyR/cADPR desensitization. Recovery from desensitization in calmodulin-depleted purified endoplasmic reticulum (microsomes) is severely impaired compared to that in crude egg homogenates. An active, soluble factor, identified as calmodulin, is required to restore the capacity of microsomes to recover from desensitization. Calmodulin mediates recovery in a manner that tightly parallels its time course of association with the RyR. Conversely, direct measurement of calmodulin binding to microsomes reveals a loss of specific binding during cADPR, but not IP(3), desensitization. Our results support a mechanism in which cycles of calmodulin dissociation and reassociation to an endoplasmic reticulum protein, most likely the RyR itself, mediate RyR/cADPR desensitization and resensitization, respectively.