Chemosensory pathways in the capitate tentacles of the hydroid Cladonema

The introduction of an extract of Artemia into the sea water bathing tentacles from the hydroid Cladonema triggers a burst of electrical activity that can be recorded intracellularly from cnidocytes in the capitate tentacles. These bursts, which are composed of a variety of events, including action potentials and EPSPs, are Ca²⁺ dependent, and are abolished by pretreatment with NiCl₂, suggesting that voltage-gated Ca²⁺ channels are involved in their generation or transmission. Intracellular injection of Lucifer Yellow and recordings from pairs of cnidocytes reveal that the cnidocytes are electrically coupled to one another, but that they are not uncoupled by heptanol. The role of these chemosensory pathways in priming the cnidocytes for discharge is discussed.     

Evidence for a common pattern of peptidergic innervation of cnidocytes

Tentacles from representatives of all four classes of the phylum Cnidaria were examined using antibodies against the neuropeptides FMRFamide and RFamide to reveal the organization of neurons and nerve nets associated with cnidocytes. The tentacles of all species examined contained FMRFamide- or RFamide-immunoreactive neurons, in varying densities. In representatives from the Scyphozoa, Hydrozoa, and Cubozoa, the FMRFamide-immunoreactive neurons formed plexuses at the base of the cnidocyte assemblages; in anthozoans, the absence of discrete assemblies of cnidocytes precluded visual co-localization of cnidocytes and immunoreactive neurons. In all four classes, immunoreactive sensory cells connected these peptidergic nerve nets to the surface of the tentacle. These findings suggest that members of all four cnidarian classes share a common organizational pattern, and it is proposed that this peptidergic innervation may be involved in the chemosensory regulation of cnidocyte discharge.     

Cnidocyte discharge is regulated by light and opsin-mediated phototransduction

Background

Cnidocytes, the eponymous cell type of the Cnidaria, facilitate both sensory and secretory functions and are among the most complex animal cell types known. In addition to their structural complexity, cnidocytes display complex sensory attributes, integrating both chemical and mechanical cues from the environment into their discharge behavior. Despite more than a century of work aimed at understanding the sensory biology of cnidocytes, the specific sensory receptor genes that regulate their function remain unknown.

Results

Here we report that light also regulates cnidocyte function. We show that non-cnidocyte neurons located in battery complexes of the freshwater polyp Hydra magnipapillata specifically express opsin, cyclic nucleotide gated (CNG) ion channel and arrestin, which are all known components of bilaterian phototransduction cascades. We infer from behavioral trials that different light intensities elicit significant effects on cnidocyte discharge propensity. Harpoon-like stenotele cnidocytes show a pronounced diminution of discharge behavior under bright light conditions as compared to dim light. Further, we show that suppression of firing by bright light is ablated by cis-diltiazem, a specific inhibitor of CNG ion channels.

Conclusions

Our results implicate an ancient opsin-mediated phototransduction pathway and a previously unknown layer of sensory complexity in the control of cnidocyte discharge. These findings also suggest a molecular mechanism for the regulation of other cnidarian behaviors that involve both photosensitivity and cnidocyte function, including diurnal feeding repertoires and/or substrate-based locomotion. More broadly, our findings highlight one novel, non-visual function for opsin-mediated phototransduction in a cnidarian, the origins of which might have preceded the evolution of cnidarian eyes.     

Lack of correlation between the amplitudes of TRP channel-mediated responses to weak and strong stimuli in intracellular Ca imaging experiments

It is often observed in intracellular Ca²⁺ imaging experiments that the amplitudes of the Ca²⁺ signals elicited by newly characterized TRP agonists do not correlate with the amplitudes of the responses evoked subsequently by a specific potent agonist. We investigated this rather controversial phenomenon by first testing whether it is inherent to the comparison of the effects of weak and strong stimuli. Using five well-characterized TRP channel agonists in commonly used heterologous expression systems we found that the correlation between the amplitudes of the Ca²⁺ signals triggered by two sequentially applied stimuli is only high when both stimuli are strong. Using mathematical simulations of intracellular Ca²⁺ dynamics we illustrate that the innate heterogeneity in expression and functional properties of Ca²⁺ extrusion (e.g. plasma membrane Ca²⁺ ATPase) and influx (TRP channels) pathways across a cellular population is a sufficient condition for low correlation between the amplitude of Ca²⁺ signals elicited by weak and strong stimuli. Taken together, our data demonstrate that this phenomenon is an expected outcome of intracellular Ca²⁺ imaging experiments that cannot be taken as evidence for lack of specificity of low-efficacy stimuli, or as an indicator of the need of other cellular components for channel stimulation.     

Rhythmic sensitization of nematocyst discharge in response to vibrational stimuli

Sea anemones capture prey by discharging nematocysts and other cnidae. Discharge of microbasic p-mastigophore (mpm) nematocysts is regulated in part by hair bundle mechanoreceptors on tentacles arising from multicellular complexes consisting of supporting cells and a sensory neuron. Anemone hair bundles detect movements of prey and then sensitize cnidocytes (cnida-containing cells) to discharge mpm nematocysts in response to contact between the prey and tentacle. Data from a simple bioassay based on counting nematocysts discharged into test probes, indicate that approximately twice as many nematocysts discharge into test probes touched to tentacles after sensitization than before sensitization. We here report that sub-second bursts of vibrational stimuli at key frequencies (51, 55, 65, or 74 Hz; Watson GM, Mire P, Hudson RR. 1998. J Exp Zool 281:582-593) sensitize discharge for at least 90 sec. Very few complete cycles of vibration are sufficient to sensitize discharge. However, as the number of cycles of vibration is increased, discharge is sensitized in rhythmic patterns. Computer analysis of the data by fast Fourier transforms indicates discharge to vibrations at 65 Hz is sensitized every 6.75 cycles. At 51 Hz discharge is sensitized every 2.00 cycles. At 74 Hz, discharge is sensitized in a polyrhythm occurring every 4.26, 3.76, 2.46, and 2. 10 cycles, respectively. At 55 Hz, discharge is sensitized in a polyrhythm occurring every 6.09, 3.20, 2.91, and 2.0 cycles, respectively. Apparently, cells in the neuronal pathway interconnecting anemone hair bundles with cnidocytes count cycles of vibration and then sensitize discharge or not according to the tally. J. Exp. Zool. 286:262-269, 2000.     

TRPs and Ca2+ in cell death and survival

Transient Receptor Potential (TRP) family mediate the influx of monovalent and/or divalent cations into cells in response to environmental stimuli. Pharmacological or genetic manipulations of TRP channels demonstrate that TRP channels influence cell death rates, prolonging or shortening of cell survival. Due to their diverse cellular localization, TRP channels mediated Ca²⁺ influx generates distinct intracellular Ca²⁺ signals that regulate downstream pathways converging to apoptosis or survival. In this review, we summarize the accumulated knowledge focused on how TRP channel regulate cell fate and may affect different pathologies including cardiovascular, neurological, metabolic or neoplastic disorders.     

Acetic acid activates PKD1L3-PKD2L1 channel—A candidate sour taste receptor

The polycystic kidney disease (PKD) 1L3-PKD2L1 channel is a candidate sour taste receptor expressed in mammalian taste receptor cells. Various acids are reported to activate PKD channels after the removal of the acid stimuli, but little information is available on the activation of these channels by acetic acid. It was difficult to analyze the PKD channel activation by acetic acid using Ca²⁺ imaging experiments because this acid induces a transient and nonspecific response in cultured cells. Here, we developed a novel method to evaluate PKD channel activation by acetic acid. Nonspecific responses were observed only over a short period after the application of acetic acid. In contrast, PKD channel activation evoked by acetic acid as well as citric acid was detected even at a later time point. This method revealed that PKD1L3-PKD2L1 channel activation by acetic acid was pH-dependent and occurred when the ambient pH was <3.1.     

Plasma membrane stretch activates transient receptor potential vanilloid and ankyrin channels in Merkel cells from hamster buccal mucosa

Merkel cells (MCs) have been proposed to form a part of the MC-neurite complex with sensory neurons. Many transient receptor potential (TRP) channels have been identified in mammals; however, the activation properties of these channels in oral mucosal MCs remain to be clarified. We investigated the biophysical and pharmacological properties of TRP vanilloid (TRPV)-1, TRPV2, TRPV4, TRP ankyrin (TRPA)-1, and TRP melastatin (TRPM)-8 channels, which are sensitive to osmotic and mechanical stimuli by measurement of intracellular free Ca²⁺ concentration ([Ca²⁺]_i) using fura-2. We also analyzed their localization patterns through immunofluorescence. MCs showed immunoreaction for TRPV1, TRPV2, TRPV4, TRPA1, and TRPM8 channels. In the presence of extracellular Ca²⁺, the hypotonic test solution evoked Ca²⁺ influx. The [Ca²⁺]_i increases were inhibited by TRPV1, TRPV2, TRPV4, or TRPA1 channel antagonists, but not by the TRPM8 channel antagonist. Application of TRPV1, TRPV2, TRPV4, TRPA1, or TRPM8 channel selective agonists elicited transient increases in [Ca²⁺]_i only in the presence of extracellular Ca²⁺. The results indicate that membrane stretching in MCs activates TRPV1, TRPV2, TRPV4, and TRPA1 channels, that it may be involved in synaptic transmission to sensory neurons, and that MCs could contribute to the mechanosensory transduction sequence.     

The single pore residue Asp in PKD2L1 determines Ca permeation of the PKD1L3/PKD2L1 complex

The polycystic kidney disease 1-like 3 (PKD1L3)–polycystic kidney disease 2-like 1 (PKD2L1) complex functions as a Ca²⁺-permeable, non-selective cation channel that is activated by acid and its subsequent removal; this is called an off-response. In this study, we identified a single aspartic residue in PKD2L1 that is responsible for the Ca²⁺ permeation of the PKD1L3/PKD2L1 complex. Calcium imaging analysis using point mutants of negatively charged amino acids present in the putative pore regions of PKD1L3 and PKD2L1 revealed that neutralization of the aspartic residue in PKD2L1 (D523N), which is conserved among PKD2 family members, abolished Ca²⁺ permeation, despite robust cell surface expression. In contrast, neutralization of the other negatively charged residues of PKD1L3 (D2049N and E2072Q) and PKD2L1 (D525N and D530N) as well as substitution of Asp⁵²³ with a glutamate residue (D523E) had little effect on Ca²⁺ permeation properties. These results demonstrate that Asp⁵²³ in PKD2L1 is a key determinant of Ca²⁺ permeation into the PKD1L3/PKD2L1 complex and that PKD2L1 contributes to forming the pore of the PKD1L3/PKD2L1 channel.     

TRPP2 channels regulate apoptosis through the Ca2+ concentration in the endoplasmic reticulum

Ca²⁺ is an important signalling molecule that regulates multiple cellular processes, including apoptosis. Although Ca²⁺ influx through transient receptor potential (TRP) channels in the plasma membrane is known to trigger cell death, the function of intracellular TRP proteins in the regulation of Ca²⁺‐dependent signalling pathways and apoptosis has remained elusive. Here, we show that TRPP2, the ion channel mutated in autosomal dominant polycystic kidney disease (ADPKD), protects cells from apoptosis by lowering the Ca²⁺ concentration in the endoplasmic reticulum (ER). ER‐resident TRPP2 counteracts the activity of the sarcoendoplasmic Ca²⁺ ATPase by increasing the ER Ca²⁺ permeability. This results in diminished cytosolic and mitochondrial Ca²⁺ signals upon stimulation of inositol 1,4,5‐trisphosphate receptors and reduces Ca²⁺ release from the ER in response to apoptotic stimuli. Conversely, knockdown of TRPP2 in renal epithelial cells increases ER Ca²⁺ release and augments sensitivity to apoptosis. Our findings indicate an important function of ER‐resident TRPP2 in the modulation of intracellular Ca²⁺ signalling, and provide a molecular mechanism for the increased apoptosis rates in ADPKD upon loss of TRPP2 channel function.     

The cnidarian cnidocyte,a hightech cellular weaponry

The members of the phylum Cnidaria (corals, sea anemones, medusae) are all equipped with stinging cells (cnidocytes, nematocytes), which serve mainly in prey capture and defense. The secretory product of these cells is a most complicated extrusome consisting of a cyst containing a tubule and a liquid matrix. Mechanical stimulation of the cell's cnidocil apparatus by a prey or an offender leads via bioelectrical signal transduction to the explosive discharge of the cnidocyst. In stenoteles of Hydra this process, during which the tubule is everted out of the cyst, takes less than 3 msec. The forces involved are partly due to spring-like tensions stored in the collagenous structural compartment, and partly to an osmotically generated intracapsular pressure, which can amount to 150 bar (1.5 × 10⁷ Pa). The osmotic machinery depends on the presence in the cyst's matrix of inorganic cations (either K⁺, Mg²⁺ or Ca²⁺) and rare polyanions (poly-γ-L-glutamates), which, so far, have not been reported from recently evolved eukaryotes. The discharging cyst acts like a self-reloading syringe, injecting poison and other components into the target. Since the cnidocytes are incapable of regenerating their exocytosed cysts, they have to be replaced by new cells derived by differentiation from pluripotent stem cells (interstitial cells).     

X-ray crystallography of TRP channels

Transient receptor potential (TRP) ion channels are molecular sensors of a large variety of stimuli including temperature, mechanical stress, voltage, small molecules including capsaicin and menthol, and lipids such as phosphatidylinositol 4,5-bisphosphate (PIP₂). Since the same TRP channels may respond to different physical and chemical stimuli, they can serve as signal integrators. Many TRP channels are calcium permeable and contribute to Ca²⁺ homeostasis and signaling. Although the TRP channel family was discovered decades ago, only recently have the structures of many of these channels been solved, largely by cryo-electron microscopy (cryo-EM). Complimentary to cryo-EM, X-ray crystallography provides unique tools to unambiguously identify specific atoms and can be used to study ion binding in channel pores. In this review we describe crystallographic studies of the TRP channel TRPV6. The methodology used in these studies may serve as a template for future structural analyses of different types of TRP and other ion channels.     

The transient receptor potential family of ion channels

The transient receptor potential (TRP) multigene superfamily encodes integral membrane proteins that function as ion channels. Members of this family are conserved in yeast, invertebrates and vertebrates. The TRP family is subdivided into seven subfamilies: TRPC (canonical), TRPV (vanilloid), TRPM (melastatin), TRPP (polycystin), TRPML (mucolipin), TRPA (ankyrin) and TRPN (NOMPC-like); the latter is found only in invertebrates and fish. TRP ion channels are widely expressed in many different tissues and cell types, where they are involved in diverse physiological processes, such as sensation of different stimuli or ion homeostasis. Most TRPs are non-selective cation channels, only few are highly Ca²⁺ selective, some are even permeable for highly hydrated Mg²⁺ ions. This channel family shows a variety of gating mechanisms, with modes of activation ranging from ligand binding, voltage and changes in temperature to covalent modifications of nucleophilic residues. Activated TRP channels cause depolarization of the cellular membrane, which in turn activates voltage-dependent ion channels, resulting in a change of intracellular Ca²⁺ concentration; they serve as gatekeeper for transcellular transport of several cations (such as Ca²⁺ and Mg²⁺), and are required for the function of intracellular organelles (such as endosomes and lysosomes). Because of their function as intracellular Ca²⁺ release channels, they have an important regulatory role in cellular organelles. Mutations in several TRP genes have been implicated in diverse pathological states, including neurodegenerative disorders, skeletal dysplasia, kidney disorders and pain, and ongoing research may help find new therapies for treatments of related diseases.     

A Pathogenic C Terminus-truncated Polycystin-2 Mutant Enhances Receptor-activated Ca2+ Entry via Association with TRPC3 and TRPC7

Mutations in PKD2 gene result in autosomal dominant polycystic kidney disease (ADPKD). PKD2 encodes polycystin-2 (TRPP2), which is a homologue of transient receptor potential (TRP) cation channel proteins. Here we identify a novel PKD2 mutation that generates a C-terminal tail-truncated TRPP2 mutant 697fsX with a frameshift resulting in an aberrant 17-amino acid addition after glutamic acid residue 697 from a family showing mild ADPKD symptoms. When recombinantly expressed in HEK293 cells, wild-type (WT) TRPP2 localized at the endoplasmic reticulum (ER) membrane significantly enhanced Ca²⁺ release from the ER upon muscarinic acetylcholine receptor (mAChR) stimulation. In contrast, 697fsX, which showed a predominant plasma membrane localization characteristic of TRPP2 mutants with C terminus deletion, prominently increased mAChR-activated Ca²⁺ influx in cells expressing TRPC3 or TRPC7. Coimmunoprecipitation, pulldown assay, and cross-linking experiments revealed a physical association between 697fsX and TRPC3 or TRPC7. 697fsX but not WT TRPP2 elicited a depolarizing shift of reversal potentials and an enhancement of single-channel conductance indicative of altered ion-permeating pore properties of mAChR-activated currents. Importantly, in kidney epithelial LLC-PK1 cells the recombinant 679fsX construct was codistributed with native TRPC3 proteins at the apical membrane area, but the WT construct was distributed in the basolateral membrane and adjacent intracellular areas. Our results suggest that heteromeric cation channels comprised of the TRPP2 mutant and the TRPC3 or TRPC7 protein induce enhanced receptor-activated Ca²⁺ influx that may lead to dysregulated cell growth in ADPKD.     

Involvement of TRPC3 channels in calcium oscillations mediated by OX1 orexin receptors

Oscillations of intracellular Ca²⁺ provide a novel mechanism for sustained activation of cellular processes. Receptor-activated oscillations are mainly thought to occur through rhythmic IP₃-dependent store discharge. However, as shown here in HEK293 cells 1 nM orexin-A (Ox-A) acting at OX₁ receptors (OX₁R) triggered oscillatory Ca²⁺ responses, requiring external Ca²⁺. These responses were attenuated by interference with TRPC3 channel (but not TRPC1/4) function using dominant negative constructs, elevated Mg²⁺ (a blocker of many TRP channels) or inhibition of phospholipase A₂. These treatments did not affect Ca²⁺ oscillations elicited by high concentrations of Ox-A (100 nM) in the absence of external Ca²⁺. OX₁R are thus able to activate TRPC(3)-channel-dependent oscillatory responses independently of store discharge.     

Molecular Bases of Multimodal Regulation of a Fungal Transient Receptor Potential (TRP) Channel

Multimodal activation by various stimuli is a fundamental characteristic of TRP channels. We identified a fungal TRP channel, TRPGz, exhibiting activation by hyperosmolarity, temperature increase, cytosolic Ca²⁺ elevation, membrane potential, and H₂O₂ application, and thus it is expected to represent a prototypic multimodal TRP channel. TRPGz possesses a cytosolic C-terminal domain (CTD), primarily composed of intrinsically disordered regions with some regulatory modules, a putative coiled-coil region and a basic residue cluster. The CTD oligomerization mediated by the coiled-coil region is required for the hyperosmotic and temperature increase activations but not for the tetrameric channel formation or other activation modalities. In contrast, the basic cluster is responsible for general channel inhibition, by binding to phosphatidylinositol phosphates. The crystal structure of the presumed coiled-coil region revealed a tetrameric assembly in an offset spiral rather than a canonical coiled-coil. This structure underlies the observed moderate oligomerization affinity enabling the dynamic assembly and disassembly of the CTD during channel functions, which are compatible with the multimodal regulation mediated by each functional module.     

TRP channels in cell survival and cell death in normal and transformed cells

TRP channels form a superfamily of channel proteins exhibiting versatile regulatory characteristics with many channels participating in the regulation of Ca²⁺ homeostasis and influencing the cell fate. Multitude of evidence is emerging that the colocalization of TRP channels with Ca²⁺-sensing elements of specific regulatory pathways leading to either proliferation or apoptosis is what makes these channels participate in cell fate regulation and, in turn, determines the final effect of Ca²⁺ entry via the particular channel. This review focuses on the aspects of TRP channel localization and function that affect the balance between cell survival and death and how various dysregulations of these channels may lead to perturbed balance and onset of cancer.     

Age attenuates the T‐type CaV3.2‐RyR axis in vascular smooth muscle

Caveolae position Ca_V3.2 (T‐type Ca²⁺ channel encoded by the α‐3.2 subunit) sufficiently close to RyR (ryanodine receptors) for extracellular Ca²⁺ influx to trigger Ca²⁺ sparks and large‐conductance Ca²⁺‐activated K⁺ channel feedback in vascular smooth muscle. We hypothesize that this mechanism of Ca²⁺ spark generation is affected by age. Using smooth muscle cells (VSMCs) from mouse mesenteric arteries, we found that both Ca_v3.2 channel inhibition by Ni²⁺ (50 µM) and caveolae disruption by methyl‐ß‐cyclodextrin or genetic abolition of Eps15 homology domain‐containing protein (EHD2) inhibited Ca²⁺ sparks in cells from young (4 months) but not old (12 months) mice. In accordance, expression of Ca_v3.2 channel was higher in mesenteric arteries from young than old mice. Similar effects were observed for caveolae density. Using SMAKO Ca_v1.2^?/? mice, caffeine (RyR activator) and thapsigargin (Ca²⁺ transport ATPase inhibitor), we found that sufficient SR Ca²⁺ load is a prerequisite for the Ca_V3.2‐RyR axis to generate Ca²⁺ sparks. We identified a fraction of Ca²⁺ sparks in aged VSMCs, which is sensitive to the TRP channel blocker Gd³⁺ (100 µM), but insensitive to Ca_V1.2 and Ca_V3.2 channel blockade. Our data demonstrate that the VSMC Ca_V3.2‐RyR axis is down‐regulated by aging. This defective Ca_V3.2‐RyR coupling is counterbalanced by a Gd³⁺ sensitive Ca²⁺ pathway providing compensatory Ca²⁺ influx for triggering Ca²⁺ sparks in aged VSMCs.     

Activity of the yeast vacuolar TRP channel TRPY1 is inhibited by Ca2+–calmodulin binding

Transient receptor potential (TRP) cation channels, which are conserved across mammals, flies, fish, sea squirts, worms, and fungi, essentially contribute to cellular Ca²⁺ signaling. The activity of the unique TRP channel in yeast, TRP yeast channel 1 (TRPY1), relies on the vacuolar and cytoplasmic Ca²⁺ concentration. However, the mechanism(s) of Ca²⁺-dependent regulation of TRPY1 and possible contribution(s) of Ca²⁺-binding proteins are yet not well understood. Our results demonstrate a Ca²⁺-dependent binding of yeast calmodulin (CaM) to TRPY1. TRPY1 activity was increased in the cmd1–6 yeast strain, carrying a non–Ca²⁺-binding CaM mutant, compared with the parent strain expressing wt CaM (Cmd1). Expression of Cmd1 in cmd1–6 yeast rescued the wt phenotype. In addition, in human embryonic kidney 293 cells, hypertonic shock-induced TRPY1-dependent Ca²⁺ influx and Ca²⁺ release were increased by the CaM antagonist ophiobolin A. We found that coexpression of mammalian CaM impeded the activity of TRPY1 by reinforcing effects of endogenous CaM. Finally, inhibition of TRPY1 by Ca²⁺–CaM required the cytoplasmic amino acid stretch E₃₃–Y₉₂. In summary, our results show that TRPY1 is under inhibitory control of Ca²⁺–CaM and that mammalian CaM can replace yeast CaM for this inhibition. These findings add TRPY1 to the innumerable cellular proteins, which include a variety of ion channels, that use CaM as a constitutive or dissociable Ca²⁺-sensing subunit, and contribute to a better understanding of the modulatory mechanisms of Ca²⁺–CaM.     

Electrical responses to water‐soluble components of fish mucus recorded from the cnidocytes of a fish predator,Physalia physalis

Physalia physalis, the Portuguese man of war, consumes mostly fish and fish larvae. Intracellular recordings from nematocyst‐containing cells (cnidocytes) in small pieces of Physalia tentacle were used to quantify the electrical responses to diluted and filtered fish skin mucus, 1–100 x 10^‐6 M amino acids, monosaccharides, and nucleosides, and seawater, which were delivered upstream of the tissue. Seawater caused responses (one pulse only) in about 10% of the applications. Fish mucus extract elicited responses in all applications, producing 1–18 depolarizing pulses (20 mV maximum amplitude). The pulses were characteristic of post‐synaptic potentials (EPSPs). Lucifer yellow and biocytin dye injections showed that the cnidocytes were not electrically coupled. Simultaneous records from two cnidocytes following mucus applications were identical.We propose, therefore, that the chemoreceptors are not on the cnidocytes, but are probably on sensory neurons that innervate clusters of cnidocytes. A < 3,000 MW fraction of mucus elicited responses indistinguishable from whole mucus extract. Higher molecular weight fractions caused no response. The various test solutions had lower percentages of response (47–92%) and produced significantly fewer pulses than the mucus extract. We conclude that prey capture in Physalia is facilitated by chemicals present in the mucus covering their fish prey. The chemical stimuli probably sensitize the nematocysts to discharge upon mechanical stimulation.