An internal model of head kinematics predicts the influence of head orientation on reflexive eye movements.

Our sense of self-motion and self-orientation results from combining information from different sources. We hypothesize that the central nervous system (CNS) uses internal models of the laws of physics to merge cues provided by different sensory systems. Different models that include internal models have been proposed; we focus herein on that referred to as the sensory weighting model. For simplicity, we isolate the portion of the sensory weighting model that estimates head angular velocity: it includes an inverse internal model of head kinematics and an 'idiotropic' vector aligned with the main body axis. Following a post-rotatory tilt in the dark, which is a rapid tilt following a constant-velocity rotation about an earth-vertical axis, the inverse internal model is applied to conflicting vestibular signals. Consequently, the CNS computes an inaccurate estimate of head angular velocity that shifts toward alignment with an estimate of gravity. Since reflexive eye movements known as vestibulo-ocular reflexes (VOR) compensate for this estimate of head angular velocity, the model predicts that the VOR rotation axis shifts toward alignment with this estimate of gravity and that the VOR time constant depends on final head orientation. These predictions are consistent with experimental data.

Reproduction of ON-center and OFF-center self-rotations.

In self-rotation reproduction tasks, subjects appear to estimate the displacement angle and then reproduce this angle without necessarily replicating the entire temporal velocity profile. In contrast, subjects appear to reproduce the entire temporal velocity profile during linear motion stimulating the otoliths. To investigate what happens during combined rotation and translation, we investigated in darkness the central processing of vestibular cues during eccentric rotation. Controlling a centrifuge with a joystick, nine healthy subjects were asked to reproduce the angle of the previously imposed rotation. Rotations were either ON-center, or 50 cm OFF-center with inter-aural centripetal acceleration. Rotation duration was either variable (proportional to the traveled angle), or constant. We examined whether the stimulation of the otoliths during OFF-center rotation changes self-rotation reproduction, and whether rotation duration is processed differently by the nervous system with and without otolith stimulation. As postulated, the subjects indeed reproduced more closely the stimulus velocity profile when OFF-center. But the primary result is that the additional supra-threshold linear acceleration cues, measured by the otoliths, did not improve performance. More specifically, to our surprise, the ability to reproduce rotation angle degraded slightly in the presence of additional information from the otolith organs, with the linear acceleration cues appearing to interfere with the reproduction of movement duration.

Neural processing of gravito-inertial cues in humans. IV. Influence of visual rotational cues during roll optokinetic stimuli.

Sensory systems often provide ambiguous information. For example, otolith organs measure gravito-inertial force (GIF), the sum of gravitational force and inertial force due to linear acceleration. However, according to Einstein's equivalence principle, a change in gravitational force due to tilt is indistinguishable from a change in inertial force due to translation. Therefore the central nervous system (CNS) must use other sensory cues to distinguish tilt from translation. For example, the CNS might use dynamic visual cues indicating rotation to help determine the orientation of gravity (tilt). This, in turn, might influence the neural processes that estimate linear acceleration, since the CNS might estimate gravity and linear acceleration such that the difference between these estimates matches the measured GIF. Depending on specific sensory information inflow, inaccurate estimates of gravity and linear acceleration can occur. Specifically, we predict that illusory tilt caused by roll optokinetic cues should lead to a horizontal vestibuloocular reflex compensatory for an interaural estimate of linear acceleration, even in the absence of actual linear acceleration. To investigate these predictions, we measured eye movements binocularly using infrared video methods in 17 subjects during and after optokinetic stimulation about the subject's nasooccipital (roll) axis (60 degrees /s, clockwise or counterclockwise). The optokinetic stimulation was applied for 60 s followed by 30 s in darkness. We simultaneously measured subjective roll tilt using a somatosensory bar. Each subject was tested in three different orientations: upright, pitched forward 10 degrees, and pitched backward 10 degrees. Five subjects reported significant subjective roll tilt (>10 degrees ) in directions consistent with the direction of the optokinetic stimulation. In addition to torsional optokinetic nystagmus and after nystagmus, we measured a horizontal nystagmus to the right during and following clockwise (CW) stimulation and to the left during and following counterclockwise (CCW) stimulation. These measurements match predictions that subjective tilt in the absence of real tilt should induce a nonzero estimate of interaural linear acceleration and, therefore, a horizontal eye response. Furthermore, as predicted, the horizontal response in the dark was larger for Tilters (n = 5) than for Non-Tilters (n = 12).

Using sensory weighting to model the influence of canal, otolith and visual cues on spatial orientation and eye movements.

The sensory weighting model is a general model of sensory integration that consists of three processing layers. First, each sensor provides the central nervous system (CNS) with information regarding a specific physical variable. Due to sensor dynamics, this measure is only reliable for the frequency range over which the sensor is accurate. Therefore, we hypothesize that the CNS improves on the reliability of the individual sensor outside this frequency range by using information from other sensors, a process referred to as "frequency completion." Frequency completion uses internal models of sensory dynamics. This "improved" sensory signal is designated as the "sensory estimate" of the physical variable. Second, before being combined, information with different physical meanings is first transformed into a common representation; sensory estimates are converted to intermediate estimates. This conversion uses internal models of body dynamics and physical relationships. Third, several sensory systems may provide information about the same physical variable (e.g., semicircular canals and vision both measure self-rotation). Therefore, we hypothesize that the "central estimate" of a physical variable is computed as a weighted sum of all available intermediate estimates of this physical variable, a process referred to as "multicue weighted averaging." The resulting central estimate is fed back to the first two layers. The sensory weighting model is applied to three-dimensional (3D) visual-vestibular interactions and their associated eye movements and perceptual responses. The model inputs are 3D angular and translational stimuli. The sensory inputs are the 3D sensory signals coming from the semicircular canals, otolith organs, and the visual system. The angular and translational components of visual movement are assumed to be available as separate stimuli measured by the visual system using retinal slip and image deformation. In addition, both tonic ("regular") and phasic ("irregular") otolithic afferents are implemented. Whereas neither tonic nor phasic otolithic afferents distinguish gravity from linear acceleration, the model uses tonic afferents to estimate gravity and phasic afferents to estimate linear acceleration. The model outputs are the internal estimates of physical motion variables and 3D slow-phase eye movements. The model also includes a smooth pursuit module. The model matches eye responses and perceptual effects measured during various motion paradigms in darkness (e.g., centered and eccentric yaw rotation about an earth-vertical axis, yaw rotation about an earth-horizontal axis) and with visual cues (e.g., stabilized visual stimulation or optokinetic stimulation).

Neural processing of gravitoinertial cues in humans. III. Modeling tilt and translation responses.

All linear accelerometers measure gravitoinertial force, which is the sum of gravitational force (tilt) and inertial force due to linear acceleration (translation). Neural strategies must exist to elicit tilt and translation responses from this ambiguous cue. To investigate these neural processes, we developed a model of human responses and simulated a number of motion paradigms used to investigate this tilt/translation ambiguity. In this model, the separation of GIF into neural estimates of gravity and linear acceleration is accomplished via an internal model made up of three principal components: 1) the influence of rotational cues (e.g., semicircular canals) on the neural representation of gravity, 2) the resolution of gravitoinertial force into neural representations of gravity and linear acceleration, and 3) the neural representation of the dynamics of the semicircular canals. By combining these simple hypotheses within the internal model framework, the model mimics human responses to a number of different paradigms, ranging from simple paradigms, like roll tilt, to complex paradigms, like postrotational tilt and centrifugation. It is important to note that the exact same mechanisms can explain responses induced by simple movements as well as by more complex paradigms; no additional elements or hypotheses are needed to match the data obtained during more complex paradigms. Therefore these modeled response characteristics are consistent with available data and with the hypothesis that the nervous system uses internal models to estimate tilt and translation in the presence of ambiguous sensory cues.

Neural processing of gravito-inertial cues in humans. II. Influence of the semicircular canals during eccentric rotation.

All linear accelerometers, including the otolith organs, respond equivalently to gravity and linear acceleration. To investigate how the nervous system resolves this ambiguity, we measured perceived roll tilt and reflexive eye movements in humans in the dark using two different centrifugation motion paradigms (fixed radius and variable radius) combined with two different subject orientations (facing-motion and back-to-motion). In the fixed radius trials, the radius at which the subject was seated was held constant while the rotation speed was changed to yield changes in the centrifugal force. In variable radius trials, the rotation speed was held constant while the radius was varied to yield a centrifugal force that nearly duplicated that measured during the fixed radius condition. The total gravito-inertial force (GIF) measured by the otolith organs was nearly identical in the two paradigms; the primary difference was the presence (fixed radius) or absence (variable radius) of yaw rotational cues. We found that the yaw rotational cues had a large statistically significant effect on the time course of perceived tilt, demonstrating that yaw rotational cues contribute substantially to the neural processing of roll tilt. We also found that the orientation of the subject relative to the centripetal acceleration had a dramatic influence on the eye movements measured during fixed radius centrifugation. Specifically, the horizontal vestibuloocular reflex (VOR) measured in our human subjects was always greater when the subject faced the direction of motion than when the subjects had their backs toward the motion during fixed radius rotation. This difference was consistent with the presence of a horizontal translational VOR response induced by the centripetal acceleration. Most importantly, by comparing the perceptual tilt responses to the eye movement responses, we found that the translational VOR component decayed as the subjective tilt indication aligned with the tilt of the GIF. This was true for both the fixed radius and variable radius conditions even though the time course of the responses was significantly different for these two conditions. These findings are consistent with the hypothesis that the nervous system resolves the ambiguous measurements of GIF into neural estimates of gravity and linear acceleration. More generally, these findings are consistent with the hypothesis that the nervous system uses internal models to process and interpret sensory motor cues.

Neural processing of gravito-inertial cues in humans. I. Influence of the semicircular canals following post-rotatory tilt.

Sensory systems often provide ambiguous information. Integration of various sensory cues is required for the CNS to resolve sensory ambiguity and elicit appropriate responses. The vestibular system includes two types of sensors: the semicircular canals, which measure head rotation, and the otolith organs, which measure gravito-inertial force (GIF), the sum of gravitational force and inertial force due to linear acceleration. According to Einstein's equivalence principle, gravitational force is indistinguishable from inertial force due to linear acceleration. As a consequence, otolith measurements must be supplemented with other sensory information for the CNS to distinguish tilt from translation. The GIF resolution hypothesis states that the CNS estimates gravity and linear acceleration, so that the difference between estimates of gravity and linear acceleration matches the measured GIF. Both otolith and semicircular canal cues influence this estimation of gravity and linear acceleration. The GIF resolution hypothesis predicts that inaccurate estimates of both gravity and linear acceleration can occur due to central interactions of sensory cues. The existence of specific patterns of vestibuloocular reflexes (VOR) related to these inaccurate estimates can be used to test the GIF resolution hypothesis. To investigate this hypothesis, we measured eye movements during two different protocols. In one experiment, eight subjects were rotated at a constant velocity about an earth-vertical axis and then tilted 90 degrees in darkness to one of eight different evenly spaced final orientations, a so-called "dumping" protocol. Three speeds (200, 100, and 50 degrees /s) and two directions, clockwise (CW) and counterclockwise (CCW), of rotation were tested. In another experiment, four subjects were rotated at a constant velocity (200 degrees /s, CW and CCW) about an earth-horizontal axis and stopped in two different final orientations (nose-up and nose-down), a so-called "barbecue" protocol. The GIF resolution hypothesis predicts that post-rotatory horizontal VOR eye movements for both protocols should include an "induced" VOR component, compensatory to an interaural estimate of linear acceleration, even though no true interaural linear acceleration is present. The GIF resolution hypothesis accurately predicted VOR and induced VOR dependence on rotation direction, rotation speed, and head orientation. Alternative hypotheses stating that frequency segregation may discriminate tilt from translation or that the post-rotatory VOR time constant is dependent on head orientation with respect to the GIF direction did not predict the observed VOR for either experimental protocol.

Effects of static orientation upon human optokinetic afternystagmus.

"Normal" human subjects were placed in a series of 5 static orientations with respect to gravity and were asked to view an optokinetic display moving at a constant angular velocity. The axis of rotation coincided with the subject's rostro-caudal axis and produced horizontal optokinetic nystagmus and afternystagmus. Wall (1) previously reported that these optokinetic afternystagmus responses were not well characterized by parametric fits to slow component velocity. The response for nose-up, however, was larger than for nose-down. This suggested that the horizontal eye movements measured during optokinetic stimulation might include an induced linear VOR component as presented in the body of this paper. To investigate this hypothesis, another analysis of these data has been made using cumulative slow component eye position. Some subjects' responses had reversals in afternystagmus direction. These reversals were "filled in" by a zero slow component velocity. This method of analysis gives a much more consistent result across subjects and shows that, on average, responses from the nose-down horizontal (prone) orientation are greatly reduced (p < 0.05) compared to other horizontal and vertical orientations. Average responses are compared to responses predicted by a model previously used to predict successfully the responses to post-rotatory nystagmus after earth horizontal axis rotation. Ten of 11 subjects had larger responses in their supine than their prone orientation. Application of horizontal axis optokinetic afternystagmus for clinical otolith function testing, and implications for altered gravity experiments are discussed.

Humans use internal models to estimate gravity and linear acceleration.

Because sensory systems often provide ambiguous information, neural processes must exist to resolve these ambiguities. It is likely that similar neural processes are used by different sensory systems. For example, many tasks require neural processing to distinguish linear acceleration from gravity, but Einstein's equivalence principle states that all linear accelerometers must measure both linear acceleration and gravity. Here we investigate whether the brain uses internal models, defined as neural systems that mimic physical principles, to help estimate linear acceleration and gravity. Internal models may be used in motor contro, sensorimotor integration and sensory processing, but direct experimental evidence for such models is limited. To determine how humans process ambiguous gravity and linear acceleration cues, subjects were tilted after being rotated at a constant velocity about an Earth-vertical axis. We show that the eye movements evoked by this post-rotational tilt include a response component that compensates for the estimated linear acceleration even when no actual linear acceleration occurs. These measured responses are consistent with our internal model predictions that the nervous system can develop a non-zero estimate of linear acceleration even when no true linear acceleration is present.

Computation of inverse dynamics for the control of movements.

Accuracy of movements requires that the central nervous system computes approximate inverse functions of the mechanical functions of limb articulations. In vertebrates, this is known to be achieved within the cerebellar pathways, and also in the cerebral cortex of primates. A cybernetic circuit achieving this computation allows accurate simulation of fast movements of the eye or forearm. It is consistent with anatomy, and with the classical view of the cerebellum as permanently supervised by the inferior olive. The inferior olive detects over-or under-shoots of movements, and the resulting climbing fiber activity corrects ongoing movements, regulates the function of cerebellar cortex and nuclei, and sets the gains of the sensorimotor reactions.

Characterization of the effects of tebufelone on hepatic cytochromes P450 in the beagle dog.

Tebufelone (1-[3,5-bis(1,1-dimethylethyl)-4-hydroxy-phenyl]-hex-5-yne-1-one) is an investigational ditertiary butylphenol nonsteroidal anti-inflammatory drug. The purpose of the present study was to assess the effects of tebufelone on hepatocyte ultrastructure and hepatic cytochromes p450 (P450s) in the beagle dog after 2 weeks of oral administration at dose levels of 0, 5, 15, 50, and 100 mg/kg/day (N = 1/sex/dose level). Hepatic tissue was obtained at necropsy for histologic, ultrastructural, and biochemical evaluation. Hepatocellular hypertrophy was observed in only a single tebufelone-treated dog (50 mg/kg). Electron microscopic evaluation, however, revealed marked dose-dependent increases in smooth endoplasmic reticulum in all of the tebufelone treatment groups. Biochemical indicators suggested that tebufelone produced mixed effects on hepatic P450s. p-Nitroanisole O-demethylase and, to a greater extent, ethoxyresorufin O-deethylase activities were decreased with increasing tebufelone dose. The precise mechanism by which tebufelone decreased ethoxyresorufin O-deethylase activity in dogs in unknown, but it was not by competitive inhibition, P450 inactivation, or reduced CYP1A expression. Tebufelone treatment increased NADPH-dependent cytochrome c reductase, total P450, and indicators of CYP2B11 (chloramphenicol covalent binding and immunochemically determined 2B11) and CYP3A12 (erythromycin N-demethylase, triacetyloleandomycin spectral complex formation, testosterone 6 beta-hydroxylase, and immunochemically determined 3A12). The largest increase in the 2B11 and 3A12 markers occurred in the 50 or 100 mg/kg treatment groups. The greatest increase in CYP2B11 markers produced by tebufelone treatment ranged from 2- to 3-fold, whereas the increase in CYP3A12 markers ranged from 5- to 10-fold. The changes in hepatic ultrastructure and increases in CYP2B11 and CYP3A12 markers produced by tebufelone in dogs are similar to that reported for phenobarbital.

Motion sickness during off-vertical axis rotation: prediction by a model of sensory interactions and correlation with other forms of motion sickness.

Motion sickness (MS) susceptibility of 108 normal subjects was measured during off-vertical axis rotation (OVAR) as a function of angular velocity (60-180 degrees/s). The chair rotated about a longitudinal axis tilted 30 degrees with respect to gravity. For each velocity, we measured the duration of exposure necessary to evoke a moderate malaise, with a limit of 30 min. MS appeared the fastest at a rotation velocity of 105 degrees/s; higher or lower velocities were less provocative. These results are in good agreement with predictions made by Zupan et al. [in ICANN'94, Springer-Verlag, 1995] by means of a MS mathematical model derived from a model of sensory interactions [Droulez and Darlot, in Attention and Performance, Vol. 13, Lawrence Erlbaum, Hillsdale, 1989]. We also found that MS susceptibility during OVAR is positively correlated with susceptibility to other forms of MS. Since OVAR induces sensory messages very different from those induced by other provocative stimulations, this could suggest that the sensitivity of a common final vegetative locus is an important factor of the individual differences in susceptibility to MS.

Structural determinants of haloenol lactone-mediated suicide inhibition of canine myocardial calcium-independent phospholipase A2.

Haloenol lactones are potent mechanism-based inhibitors of a novel class of calcium-independent phospholipases A2 which have been implicated as the enzymic mediators of membrane dysfunction during myocardial ischemia (Hazen, S. L.; et al. J. Biol. Chem. 1991, 266, 7227-7232). Herein we demonstrate that the ring size, hydrophobic group, and cryptic electrophile in the haloenol lactone moiety are important and modifiable determinants of the inhibitory potency of haloenol lactone-mediated inhibition of calcium-independent phospholipase A2. Direct comparisons between haloenol lactone-mediated inhibition of calcium-independent phospholipase A2 and the absence of inhibition with calcium-dependent phospholipase A2 further underscore the marked differences in the catalytic strategy employed by these two classes of intracellular phospholipases A2.

Cloning and expression of a human 14-3-3 protein mediating phospholipolysis. Identification of an arachidonoyl-enzyme intermediate during catalysis.

The major phospholipase A2 activity in sheep platelets is mediated by at least three chromatographically resolvable isoforms of a 30-kDa dimeric polypeptide which are responsive to physiologic increments in calcium ion and possess a dramatic substrate selectivity (Loeb, L. A., and Gross, R. W. (1986) J. Biol. Chem. 261, 10467-10470). Herein, we describe the cloning and expression of the human equivalent of one such isoform and demonstrate that it catalyzes the cleavage of the sn-2 fatty acid of choline and ethanolamine glycerophospholipids through the formation of a stable acyl-enzyme intermediate. Transesterification of the sn-2 acyl group of phosphatidylcholine to the recombinant 30-kDa polypeptide is over 50-fold selective for arachidonic acid, is augmented by calcium ion, and results in the formation of an arachidonoyl-thioester intermediate. Homology analysis demonstrated that the polypeptide mediating this transesterification is one member of a family of proteins collectively designated as 14-3-3 proteins. These results demonstrate that at least one intracellular mammalian phospholipase A2 employs a catalytic strategy distinct from that utilized by extracellular phospholipases A2 (i.e. formation of an acyl-enzyme intermediate by nucleophilic attack versus activation of a water molecule) and that arachidonic acid in endogenous phospholipid storage depots can, in principle, be sequentially transferred through an acyl-enzyme intermediate without the prior obligatory release of free arachidonic acid.

Semisynthesis and purification of homogeneous plasmenylcholine molecular species.

Methods for the efficient use of limiting amounts of fatty acid probes in the synthesis of individual molecular species of plasmenylcholine have been developed. Plasmenylcholine molecular species were synthesized through acylation of homogeneous 1-O-(Z)-hexadec-1'-enyl-sn-glycero-3-phosphocholine utilizing fatty acid anhydrides generated in situ from combined pools of reactant and recycled fatty acids by repeated addition of small amounts (10 mol%) of N,N'-dicyclohexylcarbodiimide. The efficient generation of reactive anhydrides was accomplished through minimizing irreversible formation of N-acyl urea adducts by maintaining a persistent molar excess of fatty acid (with respect to carbodiimide) during the entire reaction time course. The synthesis of multiple different sn-2 labeled plasmenylcholine probes for utilization in fluorescence, ESR, or 2H NMR spectroscopy as well as isotopically labeled plasmenylcholines for metabolic studies has been achieved in good yield (40-50% of theoretical yield based on fatty acid) by these methods. Rapid and effective purification methods utilizing high-performance liquid chromatography were developed for both large- and small-scale purifications of individual reaction mixtures which collectively resulted in the isolation of homogeneous plasmenylcholine molecular species in high yield from limiting amounts of fatty acid probes.

Calcium is sufficient but not necessary for activation of sheep platelet cytosolic phospholipase A2.

In this study we demonstrate that: (1) although the major phospholipase A2 present in sheep platelets is activated by calcium ions, it can effectively catalyze hydrolysis of the sn-2 ester linkage in phospholipids in the absence of calcium; (2) expression of calcium-independent phospholipase A2 activity can be induced by NaCl utilizing purified (but not crude) cytosolic enzyme; and (3) calcium-independent phospholipase A2 activity is regulated by a reconstitutable cytosolic protein. Collectively, these results underscore the fundamental catalytic differences between extracellular and intracellular calcium-dependent phospholipases A2 and demonstrate that calcium is sufficient, but not necessary, for the activation of this class of intracellular phospholipases A2.

Suicide inhibition of canine myocardial cytosolic calcium-independent phospholipase A2. Mechanism-based discrimination between calcium-dependent and -independent phospholipases A2.

The majority of phospholipase A2 activity in myocardium is calcium-independent and selective for hydrolysis of plasmalogen substrate (Wolf, R. A., and Gross, R. W. (1985) J. Biol. Chem. 260, 7295-7303; Hazen, S. L., Stuppy, R. J., and Gross, R. W. (1990) J. Biol. Chem. 265, 10622-10630). Accordingly, identification of an inhibitor which selectively targets calcium-independent phospholipases A2 would facilitate elucidation of the biologic significance of this class of intracellular phospholipases. We now report that the haloenol lactone, (E)-6-(bromomethylene)tetrahydro-3-(1-naphthalenyl)-2H-pyran-2-one (Compound 1), is a potent, irreversible, mechanism-based inhibitor of myocardial calcium-independent phospholipase A2 which is greater than 1000-fold specific for inhibition of myocardial calcium-independent phospholipase A2 in comparisons with multiple calcium-dependent phospholipases A2. Mechanism-based inhibition of myocardial cytosolic calcium-independent phospholipase A2 by Compound 1 was established by demonstrating: 1) time-dependent irreversible inactivation; 2) covalent binding of [3H]Compound 1 to the purified phospholipase A2; 3) ablation of covalent binding of [3H]Compound 1 after chemical inactivation of phospholipase A2 enzymic activity; 4) identical inhibition of myocardial phospholipase A2 by Compound 1 in the absence or presence of nucleophilic scavengers; 5) Compound 1 is a substrate for myocardial calcium-independent phospholipase A2 resulting in the generation of the electrophilic alpha-bromomethyl ketone; 6) phospholipase A2 inhibition requires the in situ generation of the reactive electrophile (i.e. neither the alpha-bromomethyl ketone nor the diproteoenol lactone analog are inhibitory); and 7) concomitant attenuation of the inhibitory potency and the extent of covalent adduct formation in the presence of saturating substrate. Collectively, these results demonstrate that the haloenol lactone, Compound 1, is a substrate for, covalently binds to, and irreversibly inhibits canine myocardial cytosolic calcium-independent phospholipase A2.