Inhibition of polymorphonuclear leukocyte capping by a chemotactic factor.

Incubation of human polymorphonuclear leukocytes with colchicine and fluorescein-concanavalin A leads to the formation of a polarized cap of fluorescence not seen if cells are incubated with fluorescein-Con A along. When cells are preincubated with a chemotactic factor before colchicine treatment, the capping is inhibited in a dose-related manner. Studies with alpha-methylmannoside indicate that the caps represent extracellular fluorescein-Con A and are not areas of Con A internalization. Experiments utilizing an irreversible inhibitor of serine esterases suggest that a chemotactic factor-activated enzyme is involved in the inhibition of cap formation in the human neutrophil.     

Integrating signals in stomatal development

Stomata are specialized epidermal structures that control the exchange of water and carbon dioxide between the plant and the atmosphere. The classical developmental mechanisms that define cell fate and tissue patterning - cell lineage, cell-cell interactions and signals from a distance - are employed to make stomata and to define their density and distribution within the epidermis. Recent work has shown that two genes that are involved in stomatal pattern may encode components of a classical cell-surface-receptor-mediated signaling cascade. Additional work has suggested that signals from the overlying cuticle and the underlying mesophyll also influence stomatal pattern. These findings highlight the need for models that explain how the signals that regulate stomatal development are integrated and how they act to regulate cell polarity, the cell cycle and, ultimately, cell fate.     

Consequences of chemosensory phenomena for leukocyte chemotactic orientation

The stochastic nature of cell surface receptor-ligand binding is known to limit the accuracy of detection of chemoattractant gradients by leukocytes, thus limiting the orientation ability that is crucial to the chemotactic response in host defense. The probabilistic cell orientation model of Lauffenburger is extended here to assess the consequences of recently discovered receptor phenomena: "down-regulation" of total surface receptor number, spatial asymmetry of surface receptors, and existence of a higher-affinity receptor subpopulation. In general, a reduction in orientation accuracy is predicted by inclusion of these phenomena. An orientation signal based on a simple model of chemosensory adaptation (i.e., a spatial difference in relative receptor occupancy) is found to be functionally different from the signal suggested by an experimental correlation (i.e., a spatial difference in absolute receptor occupancy). However, in the context of receptor "signal noise," the signal based on adaptation yields predictions in better qualitative agreement with the experimental orientation data of Zigmond. From this cell orientation model we can estimate the effective time-averaging period required for noise diminution to a level allowing orientation predictions to match observed levels. This time-averaging period presumably reflects the time constant for receptor signal transduction and locomotory response.     

Developmental responses of portulaca seedlings to conflicting spectral signals

Summary Portulaca oleracea seedlings avoid growing in the direction of neighbouring plants even when they are very small or remote. The present study was designed to determine the relative effect on the development of Portulaca seedlings of light availability (i.e. the resource level) as compared with spectral composition (i.e. the signal of future competition for the resource). The plants were subjected to various intensities of photosynthetic light and red/far-red (R/FR) ratios from opposite directions. The seedlings became recumbent preferentially towards the direction with the lower FR light, even when this meant growing towards plastic that absorbed 20 times more photosynthetic light. A preference for the direction with higher photosynthetic light over lower FR was also found, though only under extreme light differences. The response of the seedlings was not absolute: the orientation chosen depended on the light received from other alternative directions.     

Consequences of chemosensory phenomena for leukocyte chemotactic orientation

The stochastic nature of cell surface receptor-ligand binding is known to limit the accuracy of detection of chemoattractant gradients by leukocytes (11, 12), thus limiting the orientation ability that is crucial to the chemotactic response in host defense. The probabilistic cell orientation model of Lauffenburger (11) is extended here to assess the consequences of recently discovered receptor phenomena: “down-regulation” of total surface receptor number, spatial asymmetry of surface receptors, and existence of a higher-affinity receptor subpopulation. In general, a reduction in orientation accuracy is predicted by inclusion of these phenomena. An orientation signal based on a simple model of chemosensory adaptation (i.e., a spatial difference inrelative receptor occupancy) is found to be functionally different from the signal suggested by an experimental correlation (i.e., a spatial difference inabsolute receptor occupancy). However, in the context of receptor “signal noise,” the signal based on adaptation yields predictions in better qualitative agreement with the experimental orientation data of Zigmond (10). From this cell orientation model we can estimate the effective timeaveraging period required for noise diminution to a level allowing orientation predictions to match observed levels. This time-averaging period presumably reflects the time constant for receptor signal transduction and locomotory response.     

Pharmacological evidence is consistent with a prominent role of spatial memory in complex navigation

The ability to learn about the spatial environment plays an important role in navigation, migration, dispersal, and foraging. However, our understanding of both the role of cognition in the development of navigation strategies and the mechanisms underlying these strategies is limited. We tested the hypothesis that complex navigation is facilitated by spatial memory in a population of Chrysemys picta that navigate with extreme precision (±3.5 m) using specific routes that must be learned prior to age three. We used scopolamine, a muscarinic acetylcholine receptor antagonist, to manipulate the cognitive spatial abilities of free-living turtles during naturally occurring overland movements. Experienced adults treated with scopolamine diverted markedly from their precise navigation routes. Naive juveniles lacking experience (and memory) were not affected by scopolamine, and thereby served as controls for perceptual or non-spatial cognitive processes associated with navigation. Further, neither adult nor juvenile movement was affected by methylscopolamine, a form of scopolamine that does not cross the blood–brain barrier, a control for the peripheral effects of scopolamine. Together, these results are consistent with a role of spatial cognition in complex navigation and highlight a cellular mechanism that might underlie spatial cognition. Overall, our findings expand our understanding of the development of complex cognitive abilities of vertebrates and the neurological mechanisms of navigation.     

Influence of external concentration fluctuations on leukocyte chemotactic orientation

The quantitative dependence of leukocyte chemotactic orientation on imprecision in the measurement of chemoattractant concentrations from thermal fluctuations is analyzed. First, a mathematical model relating orientation to differences in receptor occupancy across cell dimensions is developed. This is then coupled with an extension of Berg and Purcell's analysis (1) of the precision of attractant concentration measurements by means of receptor occupancy. Our results show that thermal fluctuations in external concentrations can limit the accuracy of orientation, unless the measurement noise is reduced by averaging the measurements over a period of time. Comparison of our model predictions to experimental orientation data suggests that leukocytes do overcome this limitation, and allows estimation of the time-averaging period necessary to do so. For the orientation observed in a visual bridge assay by Zigmond (2) using the attractant peptide FNLLP, we estimate that receptor occupancy measurements for spatial comparison across cell dimensions must be averaged for a few minutes. Otherwise, the fluctuations in the attractant concentration near the cell will be too great to allow the observed degree of orientation. Our analysis also suggests that the ratio of signal-to-signal noise does not adequately characterize orientation accuracy. Accurate orientation can, in some situations, occur when this ratio is substantially less than unity; in other situations, a ratio much greater than unity is required for accurate orientation.     

Consequences of chemotactic peptide receptor modulation for leukocyte orientation

To exhibit chemotaxis, the orientation of locomotion along a chemical gradient cells sense differences in concentrations of a chemotactic factor by detecting some difference in the occupancy of their chemotactic receptors. Thus chemotaxis is sensitive to the number of receptors present and might be used to evaluate the consequences of receptor down-regulation. The ability of rabbit peritoneal polymorphonuclear leukocytes (PMNs) to orient to a standard gradient at various concentrations of N-formylnorleucylleucylphenylalanine (FNLLP) was examined. The observed orientation was compared to that expected if the directional signal were proportional to a difference in the absolute number or the fractional number of receptors occupied. The receptor occupancy in varying gradients was calculated from the binding constant of FNLLP, 2 X 10(-8) M (Zigmond and Sullivan, 1979, J. Cell Biol. 82:517-527), and the receptor number (a) present initially or (b) present after down-regulation (Sullivan and Zigmond, 1980, J. Cell Biol. 85:703-711). The observed concentration dependence of cell orientation is similar to the change in the number of receptors occupied, the receptor number being corrected for down-regulated cells. The net effect of receptor loss appears to be a decreased sensitivity to gradients at high concentrations of peptide.     

Integrating cytosolic calcium signals into mitochondrial metabolic responses.

Stimulation of hepatocytes with vasopressin evokes increases in cytosolic free Ca2+ ([Ca2+]c) that are relayed into the mitochondria, where the resulting mitochondrial Ca2+ ([Ca2+]m) increase regulates intramitochondrial Ca2+-sensitive targets. To understand how mitochondria integrate the [Ca2+]c signals into a final metabolic response, we stimulated hepatocytes with high vasopressin doses that generate a sustained increase in [Ca2+]c. This elicited a synchronous, single spike of [Ca2+]m and consequent NAD(P)H formation, which could be related to changes in the activity state of pyruvate dehydrogenase (PDH) measured in parallel. The vasopressin-induced [Ca2+]m spike evoked a transient increase in NAD(P)H that persisted longer than the [Ca2+]m increase. In contrast, PDH activity increased biphasically, with an initial rapid phase accompanying the rise in [Ca2+]m, followed by a sustained secondary activation phase associated with a decline in cellular ATP. The decline of NAD(P)H in the face of elevated PDH activity occurred as a result of respiratory chain activation, which was also manifest in a calcium-dependent increase in the membrane potential and pH gradient components of the proton motive force (PMF). This is the first direct demonstration that Ca2+-mobilizing hormones increase the PMF in intact cells. Thus, Ca2+ plays an important role in signal transduction from cytosol to mitochondria, with a single [Ca2+]m spike evoking a complex series of changes to activate mitochondrial oxidative metabolism.     

Mutants in transmission of chemotactic signals from two independent receptors of E. coli.

We have characterized chemotactic mutants of E. coli that appear to be defective in a common linkage of two independent receptors to the central chemotactic components. The mutants do not respond to gradients of ribose or galactose and thus are called trg (taxis to ribose and galactose), after Ordal and Adler (1974b). These trg mutants are indistinguishable from their parent in tactic response to other attractants, swimming pattern, growth rates, and transport of ribose and galactose. The mutant cells contain the usual amounts of ribose and galactose receptors, and those proteins function normally in their other role, transport of their respective ligands. The mutations, generated by insertion of translocatable drug-resistance elements (transposons)8 are located near 31 min on the map of the E. coli chromosome, a locus far removed from the genes coding for the ribose and galactose receptors. Trg mutants do not resemble either specific receptor mutants or che mutants. The nature of the requirement for the trg product in the response to ribose and galactose is not defined, but evidence for interference of tactic signals from the ribose and galactose receptors (Strange and Koshland, 1976) supports the idea that the product functions directly in the transmission of tactic signals from the two receptors to the flagella.     

The role of phospholipases and phospholipid-derived signals in cell activation.

The complexity of receptor-regulated breakdown and modification of phospholipids continues to grow. New developments extend our concepts of signalling enzymes and possible messengers.     

Amino acid efflux in response to chemotactic and osmotic signals in Bacillus subtilis.

We observed a large efflux of nonvolatile radioactivity from Bacillus subtilis in response to the addition of 31 mM butyrate or the withdrawal of 0.1 M aspartate in a flow assay. The major nonvolatile components effluxed were methionine, proline, histidine, and lysine. In studies of the release of volatile radioactivity in chemotaxis by B. subtilis cells that had been labeled with [3H]methionine, the breakdown of methionine to methanethiol can contribute substantially to the volatile radioactivity in fractions following addition of 0.1 M aspartate. However, methanol was confirmed to be released after aspartate addition and, in lesser quantities, after aspartate withdrawal. Methanol and methanethiol were positively identified by derivitization with 3,5-dinitro-benzoylchloride. Amino acid efflux but not methanol release was observed in response to 0.1 M aspartate stimulation of a cheR mutant of B. subtilis that lacks the chemotaxis methylesterase. The amino acid efflux could be reproduced by withdrawal of 0.1 M NaCl, 0.2 M sucrose, or 0.2 M xylitol and is probably the result of changes in osmolarity. Chemotaxis to 10 mM alanine or 10 mM proline resulted in methanol release but not efflux of amino acids. In behavioral studies, B. subtilis tumbled for 16 to 18 s in response to a 200 mosM upshift and for 14 s after a 20 mosM downshift in osmolarity when the bacteria were in perfusion buffer (40 mosM). The pattern of methanol release was similar to that observed in chemotaxis. This is consistent with osmotaxis in B. subtilis away from an increase or decrease in the osmolarity of the incubation medium. The release of methanol suggests that osmotaxis is correlated with methylation of a methyl-accepting chemotaxis protein.     

Insulin, at physiological concentrations, enhances the polymorphonuclear leukocyte chemotactic properties.

We investigated the influence of insulin on human polymorphonuclear leukocyte (PMN) chemotaxis induced by mediators in a microchamber assay. Insulin increased, with a dose-response relationship, chemotaxis induced by formyl-methionyl-leucyl-phenylalanine, calcium ionophore and phorbol-miristyl acetate (p = 0.0057, p = 0.0001 and p = 0.0215, respectively). The hormone effect was present also at the physiological concentration of 40 microU/ml. Our data show that insulin affects PMN activity in normal subjects and therefore support the hypothesis that insulin deficiency may be responsible for the impaired PMN function observed in diabetic patients in poor metabolic control.     

Sensory adaptation of Dictyostelium discoideum cells to chemotactic signals

Postvegetative Dictyostelium discoideum cells react chemotactically to gradients of cAMP, folic acid, and pterin. In the presence of a constant concentration of 10(-5) M cAMP cells move at random. They still are able to respond to superimposed gradients of cAMP, although the response is less efficient than without the high background level of cAMP. Cells which are accommodated to 10(-5) M cAMP do not react to a gradient of cAMP if the mean cAMP concentration is decreasing with time. This indicates the involvement of adaptation in the detection of chemotactic gradients: cells adapt to the mean concentration of chemoattractant and respond to positive deviations from the mean concentration. Cells adapted to high cAMP concentrations react normally to gradients of folic acid or pterin. Adaptation to one of these compounds does not affect the response to the other attractants. This suggests that cAMP, folic acid, and pterin are detected by different receptors, and that adaptation is localized at a step in the transduction process before the signals from these receptors coincide into one pathway. I discuss the implications of adaptation for chemotaxis and cell aggregation.     

The preferential human mononuclear leukocyte chemotactic activity of the substituent tetrapeptides of angiotensin II

The amino- and carboxy-terminal substituent tetrapeptides of angiotensin II, Asp-Arg-Val-Tyr and Ile-His-Pro-Phe, elicit substantial human mononuclear leukocyte chemotactic responses $\text{in}\text{vitro}$ that attain maximal levels at tetrapeptide concentrations of 3 × 10^?8 M and 3 × 10^?7 M, respectively. In contrast, the angiotensin II-derived tetrapeptides evoke only marginal human neutrophil chemotactic responses. Amino acid deletions or substitutions that alter the properties of the tetrapeptides, reduce their chemotactic potency and activity. Limited proteolytic cleavage of angiotensin II thus may convert a pathway with predominantly humoral effects to a source of mediators that regulate cellular immunity and chronic inflammatory responses.     

The oligopeptide chemotactic factor receptor on human polymorphonuclear leukocyte membranes exists in two affinity states

The binding of the chemoattractant N-formyl-methionylleucyl-[³H]phenylalanine to intact polymorphonuclear leukocytes and membrane preparations was analyzed by computer methods. Whole viable cells bind the chemoattractant with a single dissociation constant (K_D) of 22.3 ± 2.4 nM and contain an average of 55,000 receptors percell. In contrast, the binding data using membrane preparations were consistent with the presence of two classes of binding sites with average K_Ds of 0.53 ± 0.01 nM and 24.4 ± 1.2 nM. The high affinity receptors accounted for ca. 25% of the binding sites. Increasing the receptor occupancy did not affect the rate of dissociation of the ligand-receptor complex thus negative cooperativity is not a likely explanation for the complex binding isotherms. On the other hand, the dissociation kinetics did agree with the two affinity receptor model.     

Integrating individual search and navigation behaviors in mechanistic movement models

Understanding complex movement behaviors via mechanistic models is one key challenge in movement ecology. We built a theoretical simulation model using evolutionarily trained artificial neural networks (ANNs) wherein individuals evolve movement behaviors in response to resource landscapes on which they search and navigate. We distinguished among non-oriented movements in response to proximate stimuli, oriented movements utilizing perceptual cues from distant targets, and memory mechanisms that assume prior knowledge of a target??s location and then tested the relevance of these three movement behaviors in relation to size of resource patches, predictability of resource landscapes, and the occurrence of movement barriers. Individuals were more efficient in locating resources under larger patch sizes and predictable landscapes when memory was advantageous. However, memory was also frequently used in unpredictable landscapes with intermediate patch sizes to systematically search the entire spatial domain, and because of this, we suggest that memory may be important in explaining super-diffusion observed in many empirical studies. The sudden imposition of movement barriers had the greatest effect under predictable landscapes and temporarily eliminated the benefits of memory. Overall, we demonstrate how movement behaviors that are linked to certain cognitive abilities can be represented by state variables in ANNs and how, by altering these state variables, the relevance of different behaviors under different spatiotemporal resource dynamics can be tested. If adapted to fit empirical movement paths, methods described here could help reveal behavioral mechanisms of real animals and predict effects of anthropogenic landscape changes on animal movement.