Extending in vitro conditioning in Aplysia to analyze operant and classical processes in the same preparation.

Operant and classical conditioning are major processes shaping behavioral responses in all animals. Although the understanding of the mechanisms of classical conditioning has expanded significantly, the understanding of the mechanisms of operant conditioning is more limited. Recent developments in Aplysia are helping to narrow the gap in the level of understanding between operant and classical conditioning, and have raised the possibility of studying the neuronal processes underlying the interaction of operant and classical components in a relatively complex learning task. In the present study, we describe a first step toward realizing this goal, by developing a single in vitro preparation in which both operant and classical conditioning can be studied concurrently. The new paradigm reproduced previously published results, even under more conservative and homogenous selection criteria and tonic stimulation regime. Moreover, the observed learning was resistant to delay, shortening, and signaling of reinforcement.

In vitro analog of classical conditioning of feeding behavior in aplysia.

The feeding behavior of Aplysia californica can be classically conditioned using tactile stimulation of the lips as a conditioned stimulus (CS) and food as an unconditioned stimulus (US). Moreover, several neural correlates of classical conditioning have been identified. The present study extended previous work by developing an in vitro analog of classical conditioning and by investigating pairing-specific changes in neuronal and synaptic properties. The preparation consisted of the isolated cerebral and buccal ganglia. Electrical stimulation of a lip nerve (AT4) and a branch of the esophageal nerve (En2) served as the CS and US, respectively. Three protocols were used: paired, unpaired, and US alone. Only the paired protocol produced a significant increase in CS-evoked fictive feeding. At the cellular level, classical conditioning enhanced the magnitude of the CS-evoked synaptic input to pattern-initiating neuron B31/32. In addition, paired training enhanced both the magnitude of the CS-evoked synaptic input and the CS-evoked spike activity in command-like neuron CBI-2. The in vitro analog of classical conditioning reproduced all of the cellular changes that previously were identified following behavioral conditioning and has led to the identification of several new learning-related neural changes. In addition, the pairing-specific enhancement of the CS response in CBI-2 indicates that some aspects of associative plasticity may occur at the level of the cerebral sensory neurons.

Altered processing of contextual information during fear extinction in PTSD: an fMRI study.

Medial prefrontal cortical areas have been hypothesized to underlie altered contextual processing in posttraumatic stress disorder (PTSD). We investigated brain signaling of contextual information in this disorder. Eighteen PTSD subjects and 16 healthy trauma-exposed subjects underwent a two-day fear conditioning and extinction paradigm. On day 1, within visual context A, a conditioned stimulus (CS) was followed 60% of the time by an electric shock (conditioning). The conditioned response was then extinguished (extinction learning) in context B. On day 2, recall of the extinction memory was tested in context B. Skin conductance response (SCR) and functional magnetic resonance imaging (fMRI) data were collected during context presentations. There were no SCR group differences in any context presentation. Concerning fMRI data, during late conditioning, when context A signaled danger, PTSD subjects showed dorsal anterior cingulate cortical (dACC) hyperactivation. During early extinction, when context B had not yet fully acquired signal value for safety, PTSD subjects still showed dACC hyperactivation. During late extinction, when context B had come to signal safety, they showed ventromedial prefrontal cortex (vmPFC) hypoactivation. During early extinction recall, when context B signaled safety, they showed both vmPFC hypoactivation and dACC hyperactivation. These findings suggest that PTSD subjects show alterations in the processing of contextual information related to danger and safety. This impairment is manifest even prior to a physiologically-measured, cue-elicited fear response, and characterized by hypoactivation in vmPFC and hyperactivation in dACC.

Molecular mechanisms of classical fear conditioning : GABAergic factors and their role in fear-related network activities

Magdeburg, Univ., Fak. für Naturwiss., Diss., 2010

Unconditioned responses and functional fear networks in human classical conditioning.

Human imaging studies examining fear conditioning have mainly focused on the neural responses to conditioned cues. In contrast, the neural basis of the unconditioned response and the mechanisms by which fear modulates inter-regional functional coupling have received limited attention. We examined the neural responses to an unconditioned stimulus using a partial-reinforcement fear conditioning paradigm and functional MRI. The analysis focused on: (1) the effects of an unconditioned stimulus (an electric shock) that was either expected and actually delivered, or expected but not delivered, and (2) on how related brain activity changed across conditioning trials, and (3) how shock expectation influenced inter-regional coupling within the fear network. We found that: (1) the delivery of the shock engaged the red nucleus, amygdale, dorsal striatum, insula, somatosensory and cingulate cortices, (2) when the shock was expected but not delivered, only the red nucleus, the anterior insular and dorsal anterior cingulate cortices showed activity increases that were sustained across trials, and (3) psycho-physiological interaction analysis demonstrated that fear led to increased red nucleus coupling to insula but decreased hippocampus coupling to the red nucleus, thalamus and cerebellum. The hippocampus and the anterior insula may serve as hubs facilitating the switch between engagement of a defensive immediate fear network and a resting network.

Role of the hippocampus in contextual memory after classical aversive conditioning in pigeons (C. livia)

We investigated the effects of hippocampal lesions with ibotenic acid (IBO) on the memory of the sound-context-shock association during reexposure to the conditioning context. Twenty-nine adult pigeons were assigned to a non-lesioned control group (CG, N = 7), a sham-lesioned group (SG, N = 7), a hippocampus-lesioned experimental group (EG, N = 7), and to an unpaired nonlesioned group (tone-alone exposure) (NG, N = 8). All pigeons were submitted to a 20-min session in the conditioning chamber with three associations of sound (1000 Hz, 85 dB, 1 s) and shock (10 mA, 1 s). Experimental and sham lesions were performed 24 h later (EG and SG) when EG birds received three bilateral injections (anteroposterior (A), 4.5, 5.25 and 7.0) of IBO (1 µl and 1 µg/µl) and SG received one bilateral injection (A, 5.25) of PBS. The animals were reexposed to the training context 5 days after the lesion. Behavior was videotaped for 20 min and analyzed at 30-s intervals. A significantly higher percent rating of immobility was observed for CG (median, 95.1; range, 79.2 to 100.0) and SG (median, 90.0; range, 69.6 to 95.0) compared to EG (median, 11.62; range, 3.83 to 50.1) and NG (median, 7.33; range, 6.2 to 28.1) (P<0.001) in the training context. These results suggest impairment of contextual fear in birds who received lesions one day after conditioning and a role for the hippocampus in the modulation of emotional aversive memories in pigeons.

Cerebellar cortex involvement in classical eyelid conditioning

A model for cerebellar involvement in motor learning was tested using classical eyelid conditioning in the rabbit. Briefly, we assume that modifications of the strength of granule cell synapses at Purkinje cells in the cerebellar cortex and mossy fiber (MF) synapses at cerebellar interpositus nuclei are responsible for the acquisition, adaptively-timed expression, and extinction of conditioned eyelid responses (CRs). A corollary of these assumptions is that the cerebellar cortex is necessary for acquisition and extinction. This model also suggests a mechanism whereby the cerebellar cortex can discriminate different times during a conditioned stimulus (CS) and thus mediate the learned timing of CRs. Therefore, experiments were done to determine the role of the cerebellar cortex in the timing, extinction, and acquisition of CRs. Lesions of the cerebellar cortex that included the anterior lobe disrupted the learned timing of CRs such that they occurred at extremely short latencies. Stimulation of MFs in the middle cerebellar peduncle as the CS could support differently timed CRs in the same animal. These data indicate that synaptic plasticity in the cerebellar cortex mediates the learned timing of CRs. These short-latency CRs which resulted from anterior lobe damage did not extinguish, while CRs in animals receiving lesions which did not include the anterior lobe extinguished normally. Preliminary data suggests that lesions of the anterior lobe which produce short-latency responses prevent the acquisition of CRs to a novel CS. These findings indicate that the anterior lobe of cerebellar cortex is necessary for eyelid conditioning. The involvement of the anterior lobe in eyelid conditioning has not been previously reported, however, the anterior lobe has generally been spared in lesion studies examining cerebellar cortex involvement in eyelid conditioning due to its relatively inaccessible location. The observation that the anterior lobe of the cerebellar cortex is not always required for the basic expression of CRs, but is necessary for response timing, extinction, and acquisition, is consistent with the hypothesis that eyelid conditioning can involve plasticity in both the cerebellar cortex and interpositus nucleus and that plasticity in the nucleus is controlled by Purkinje cell activity. ^

Behavioral and cellular analysis of classical conditioning of feeding behavior in Aplysia

In classical conditioning, an associative form of learning, animals learn to associate two stimuli. Cellular and molecular mechanisms for the induction and consolidation of associative learning and memory at the level of single cells and synaptic connections have been studied in both vertebrate and invertebrate animals. The majority of studies, however, relied on aversive stimuli to induce learning. This bias may limit the extent to which identified mechanisms generalize to other forms of associative learning and memory, such as appetitive forms. The goal of the present study was to develop a classical conditioning procedure for the marine mollusk Aplysia californica using appetitive reinforcement, and to analyze associative learning using behavioral and electrophysiological techniques. ^ Using tactile stimulation of the lips as the conditional stimulus (CS) and food as the unconditional stimulus (US) a training protocol was developed that reliably induced classical conditioning of feeding behavior. Memory persisted for at least 24 hours. The gross organization of reinforcement-mediating pathways was analyzed in additional behavioral experiments. Moreover, neurophysiological correlates of classical conditioning were identified and characterized in an in vitro preparation containing the circuitry for feeding behavior. In vitro stimulation of a nerve (AT4) that may mediate the CS during training, resulted in a greater number of buccal motor patterns (BMPs) in brains from conditioned animals, as compared to control animals. The majority of these BMPs were ingestion-like, consistent with the increased number of bites in response to the CS after classical conditioning. Moreover, classical conditioning correlated with increased excitatory synaptic input to BMP-initiating neuron B31/32, in response to stimulation of AT 4, as compared to controls. The expression of the correlates of classical conditioning identified in this study was specific to stimulation of AT 4, which is consistent the stimulus specificity that is characteristic for classical conditioning. ^ The identification of cellular correlates of classical conditioning documented here provides the basis for future, more detailed analyses of an appetitive form of associative learning and memory, that may extend the working knowledge of the cellular and molecular mechanisms for associative plasticity in general. ^

Cellular mechanisms of operant and classical conditioning

The ability to associate a predictive stimulus with a subsequent salient event (i.e., classical conditioning) and the ability to associate an expressed behavior with the consequences (i.e., operant conditioning) allow for a predictive understanding of a changing environment. Although they are operationally distinct, there has been considerable debate whether at some fundamental level classical and operant conditioning are mechanistically distinct or similar. Feeding behavior of Aplysia (i.e., biting) was chosen as the model system and was successfully conditioned with appetitive forms of both operant and classical conditioning. The neuronal circuitry responsible for feeding is well understood and is suitable for cellular analyses, thus providing for a mechanistic comparison between these two forms of associative learning. ^ Neuron B51 is part of the feeding circuitry of Aplysia and is critical for the expression of ingestive behaviors. B51 also is a locus of plasticity following both operant and classical conditioning. Both in vivo and in vitro operant conditioning increased the input resistance and the excitability of B51. No pairing-specific changes in the input resistance were observed following both in vivo and in vitro classical conditioning. However, classical conditioning decreased the excitability of B51. Thus, both operant and classical conditioning modified the threshold level for activation of neuron B51, but in opposite directions, revealing key differences in the cellular mechanisms underlying these two forms of associative learning. ^ Next, the cellular mechanisms underlying operant conditioning were investigated in more detail using a single-cell analogue. The single-cell analogue successfully recapitulated the previous in vivo and in vitro operant conditioning results by increasing the input resistance and the excitability of B51. Both PKA and PKC were necessary for operant conditioning. Dopamine appears to be the transmitter mediating the reinforcement signal in this form of conditioning. A D1 dopamine receptor antibody revealed that the D1receptor localizes to the axon hillock, which is also the region that gives the strongest response when iontophoresing dopamine. ^ The studies presented herein, thus, provide for a greater understanding of the mechanisms underlying both of these forms of associative learning and demonstrate that they likely operate through distinct cellular mechanisms. ^

Learning processes and the neural analysis of conditioning

Classical and operant conditioning principles, such as the behavioral discrepancy-derived assumption that reinforcement always selects antecedent stimulus and response relations, have been studied at the neural level, mainly by observing the strengthening of neuronal responses or synaptic connections. A review of the literature on the neural basis of behavior provided extensive scientific data that indicate a synthesis between the two conditioning processes based mainly on stimulus control in learning tasks. The resulting analysis revealed the following aspects. Dopamine acts as a behavioral discrepancy signal in the midbrain pathway of positive reinforcement, leading toward the nucleus accumbens. Dopamine modulates both types of conditioning in the Aplysia mollusk and in mammals. In vivo and in vitro mollusk preparations show convergence of both types of conditioning in the same motor neuron. Frontal cortical neurons are involved in behavioral discrimination in reversal and extinction procedures, and these neurons preferentially deliver glutamate through conditioned stimulus or discriminative stimulus pathways. Discriminative neural responses can reliably precede operant movements and can also be common to stimuli that share complex symbolic relations. The present article discusses convergent and divergent points between conditioning paradigms at the neural level of analysis to advance our knowledge on reinforcement.

Conditioned inhibition of autonomic Pavlovian conditioning in humans

The present study aimed to demonstrate conditioned inhibition of Pavlovian conditioning of autonomic responses in humans. Subjects (N = 21) were presented initially with four geometric shapes (A, B, C and D). An electric shock served as the unconditioned stimulus (US) during acquisition. Conditional stimuli lasted for 8 s and US onset coincided with CS offset. Subjects were trained with A-US, C-US, and AC-US pairings and AB alone and B alone presentations. The subsequent summation test consisted of C-US pairings and CB alone and CD alone presentations. Conditioning was evident in self-reported US expectancy and first and second interval electrodermal responses. Evidence for conditioned inhibition during the summation test was found in US expectancy and second interval electrodermal responses. (C) 1997 Elsevier Science B.V.

Conditioning of multiple-point statistics facies simulations to tomographic images

Geophysical tomography captures the spatial distribution of the underlying geophysical property at a relatively high resolution, but the tomographic images tend to be blurred representations of reality and generally fail to reproduce sharp interfaces. Such models may cause significant bias when taken as a basis for predictive flow and transport modeling and are unsuitable for uncertainty assessment. We present a methodology in which tomograms are used to condition multiple-point statistics (MPS) simulations. A large set of geologically reasonable facies realizations and their corresponding synthetically calculated cross-hole radar tomograms are used as a training image. The training image is scanned with a direct sampling algorithm for patterns in the conditioning tomogram, while accounting for the spatially varying resolution of the tomograms. In a post-processing step, only those conditional simulations that predicted the radar traveltimes within the expected data error levels are accepted. The methodology is demonstrated on a two-facies example featuring channels and an aquifer analog of alluvial sedimentary structures with five facies. For both cases, MPS simulations exhibit the sharp interfaces and the geological patterns found in the training image. Compared to unconditioned MPS simulations, the uncertainty in transport predictions is markedly decreased for simulations conditioned to tomograms. As an improvement to other approaches relying on classical smoothness-constrained geophysical tomography, the proposed method allows for: (1) reproduction of sharp interfaces, (2) incorporation of realistic geological constraints and (3) generation of multiple realizations that enables uncertainty assessment.

Anterograde effects of a single electroconvulsive shock on inhibitory avoidance and on cued fear conditioning

A single electroconvulsive shock (ECS) or a sham ECS was administered to male 3-4-month-old Wistar rats 1, 2, and 4 h before training in an inhibitory avoidance test and in cued classical fear conditioning (measured by means of freezing time in a new environment). ECS impaired inhibitory avoidance at all times and, at 1 or 2 h before training, reduced freezing time before and after re-presentation of the ECS. These results are interpreted as a transient conditioned stimulus (CS)-induced anxiolytic or analgesic effect lasting about 2 h after a single treatment, in addition to the known amnesic effect of the stimulus. This suggests that the effect of anterograde learning impairment is demonstrated unequivocally only when the analgesic/anxiolytic effect is over (about 4 h after ECS administration) and that this impairment of learning is selective, affecting inhibitory avoidance but not classical fear conditioning to a discrete stimulus.

Frequency-adjustable positive sequence detector for power conditioning applications

This paper proposes a novel and simple positive sequence detector (PSD), which is inherently self-adjustable to fundamental frequency deviations by means of a software-based PLL (Phase Locked Loop). Since the proposed positive sequence detector is not based on Fortescue's classical decomposition and no special input filtering is needed, its dynamic response may be as fast as one fundamental cycle. The digital PLL ensures that the positive sequence components can be calculated even under distorted waveform conditions and fundamental frequency deviations. For the purpose of validating the proposed models, the positive sequence detector has been implemented in a PC-based Power Quality Monitor and experimental results illustrate its good performance. The PSD algorithm has also been evaluated in the control loop of a Series Active Filter and simulation results demonstrate its effectiveness in a closed-loop system. Moreover, considering single-phase applications, this paper also proposes a general single-phase PLL and a Fundamental Wave Detector (FWD) immune to frequency variations and waveform distortions. © 2005 IEEE.

Distance-based kriging relying on proxy simulations for inverse conditioning

Let us consider a large set of candidate parameter fields, such as hydraulic conductivity maps, on which we can run an accurate forward flow and transport simulation. We address the issue of rapidly identifying a subset of candidates whose response best match a reference response curve. In order to keep the number of calls to the accurate flow simulator computationally tractable, a recent distance-based approach relying on fast proxy simulations is revisited, and turned into a non-stationary kriging method where the covariance kernel is obtained by combining a classical kernel with the proxy. Once the accurate simulator has been run for an initial subset of parameter fields and a kriging metamodel has been inferred, the predictive distributions of misfits for the remaining parameter fields can be used as a guide to select candidate parameter fields in a sequential way. The proposed algorithm, Proxy-based Kriging for Sequential Inversion (ProKSI), relies on a variant of the Expected Improvement, a popular criterion for kriging-based global optimization. A statistical benchmark of ProKSI’s performances illustrates the efficiency and the robustness of the approach when using different kinds of proxies.