Selective hippocampal lesions disrupt a novel cue effect but fail to eliminate blocking in rabbit eyeblink conditioning

The classical conditioning task of blocking involves the adding of a novel but redundant stimulus to a previously trained stimulus. Both blocking and novelty detection are thought to involve the hippocampus. Previously, Solomon (1977) found that nonselective aspiration lesions of the hippocampal region eliminated blocking in rabbit eyeblink conditioning. We tested the effects of selective ibotenic acid lesions of the hippocampus on blocking, as well as on novelty detection, when training is switched from a tone conditioned stimulus (CS) to a compound tone-light CS in eyeblink conditioning. Selective hippocampal lesions did not eliminate blocking but did lead to a facilitation of conditioned response (CR) acquisition to the tone and to the light, but not to the tone-light compound. Selective hippocampal lesions disrupted a CR decrement observed in sham surgical controls when transferred from tone training to tone-light training. It appears that although selective hippocampal lesions do not eliminate blocking in eyeblink conditioning, they do disrupt novelty detection and may facilitate learning to a previously blocked cue.     

Fear develops to the conditioned stimulus and to the context during classical eyeblink conditioning in rats

In classical eyeblink conditioning, non-specific emotional responses to the aversive shock unconditioned stimulus (US), which are presumed to coincide with the development of fear, occur early in conditioning and precede the emergence of eyeblink responses. This twoprocess learning model was examined by concurrently measuring fear and eyeblink conditioning in the freely moving rat. Freezing served as an index of fear in animals and was measured during the inter-trial intervals in the training context and during a tone conditioned stimulus (CS) presented in a novel context. Animals that received CS-US pairings exhibited elevated levels of fear to the context and CS early in training that decreased over sessions, while eyeblink conditioned responses (CRs) developed gradually during acquisition and decreased during extinction. Random CS-US presentations produced a similar pattern of fear responses to the context and CS as paired presentations despite low eyeblink CR percentages, indicating that fear responding was decreased independent of high levels of learned eyeblink responding The results of paired training were consistent with two-process models of conditioning that postulate that early emotional responding facilitates subsequent motor learning, but measures from random control animals demonstrate that partial CS-US contingencies produce decrements in fear despite low levels of eyeblink CRs. These findings suggest, a relationship between CS-US contingency and fear levels during eyeblink conditioning, and may serve to clarify further the role that fear conditioning plays in this simple paradigm.     

Fear Develops to the Conditioned Stimulus and to the Context during Classical Eyeblink Conditioning in Rats

In classical eyeblink conditioning, non-specific emotional responses to the aversive shock unconditioned stimulus (US), which are presumed to coincide with the development of fear, occur early in conditioning and precede the emergence of eyeblink responses. This two-process learning model was examined by concurrently measuring fear and eyeblink conditioning in the freely moving rat. Freezing served as an index of fear in animals and was measured during the inter-trial intervals in the training context and during a tone conditioned stimulus (CS) presented in a novel context. Animals that received CS-US pairings exhibited elevated levels of fear to the context and CS early in training that decreased over sessions, while eyeblink conditioned responses (CRs) developed gradually during acquisition and decreased during extinction. Random CS-US presentations produced a similar pattern of fear responses to the context and CS as paired presentations despite low eyeblink CR percentages, indicating that fear responding was decreased independent of high levels of learned eyeblink responding. The results of paired training were consistent with two-process models of conditioning that postulate that early emotional responding facilitates subsequent motor learning, but measures from random control animals demonstrate that partial CS-US contingencies produce decrements in fear despite low levels of eyeblink CRs. These findings suggest a relationship between CS-US contingency and fear levels during eyeblink conditioning, and may serve to clarify further the role that fear conditioning plays in this simple paradigm.     

Hippocampal and cerebellar single-unit activity during delay and trace eyeblink conditioning in the rat

In delay eyeblink conditioning, the CS overlaps with the US and only a brainstem-cerebellar circuit is necessary for learning. In trace eyeblink conditioning, the CS ends before the US is delivered and several forebrain structures, including the hippocampus, are required for learning, in addition to a brainstem-cerebellar circuit. The interstimulus interval (ISI) between CS onset and US onset is perhaps the most important factor in classical conditioning, but studies comparing delay and trace conditioning have typically not matched these procedures in this crucial factor, so it is often difficult to determine whether results are due to differences between delay and trace or to differences in ISI. In the current study, we employed a 580-ms CS-US interval for both delay and trace conditioning and compared hippocampal CA1 activity and cerebellar interpositus nucleus activity in order to determine whether a unique signature of trace conditioning exists in patterns of single-unit activity in either structure. Long-Evans rats were chronically implanted in either CA1 or interpositus with microwire electrodes and underwent either delay eyeblink conditioning, or trace eyeblink conditioning with a 300-ms trace period between CS offset and US onset. On trials with a CR in delay conditioning, CA1 pyramidal cells showed increases in activation (relative to a pre-CS baseline) during the CS-US period in sessions 1-4 that was attenuated by sessions 5-6. In contrast, on trials with a CR in trace conditioning, CA1 pyramidal cells did not show increases in activation during the CS-US period until sessions 5-6. In sessions 5-6, increases in activation were present only to the CS and not during the trace period. For rats with interpositus electrodes, activation of interpositus neurons on CR trials was present in all sessions in both delay and trace conditioning. However, activation was greater in trace compared to delay conditioning in the first half of the CS-US interval (during the trace CS) during early sessions of conditioning and, in later sessions of conditioning, activation was greater in the second half of the CS-US interval (during the trace interval). These results suggest that the pattern of hippocampal activation that differentiates trace from delay eyeblink conditioning is a slow buildup of activation to the CS, possibly representing encoding of CS duration or discrimination of the CS from the background context. Interpositus nucleus neurons show strong modeling of the eyeblink CR regardless of paradigm but show a changing pattern across conditioning that may be due to the necessary contributions of forebrain processing to trace conditioning.     

Medial auditory thalamic input to the lateral pontine nuclei is necessary for auditory eyeblink conditioning

Auditory and visual conditioned stimulus (CS) pathways for eyeblink conditioning were investigated with reversible inactivation of the medial (MPN) or lateral (LPN) pontine nuclei. In Experiment 1, Long-Evans rats were given three phases of eyeblink conditioning. Phase 1 consisted of three training sessions with electrical stimulation of the medial auditory thalamic nuclei (MATN) paired with a periorbital shock unconditioned stimulus (US). An additional session was given with a muscimol (0.5 μL, 10 mM) or saline infusion targeting the LPN followed by a recovery session with no infusions. The same training and testing sequence was then repeated with either a tone or light CS in phases 2 and 3 (counterbalanced). Experiment 2 consisted of the same training as Experiment 1 except that muscimol or saline was infused in the MPN during the retention tests. Muscimol infusions targeting the LPN severely impaired retention of eyeblink conditioned responses (CRs) to the MATN stimulation and tone CSs but only partially reduced CR percentage to the light CS. Muscimol infusions that targeted the MPN had a larger effect on CR retention to the light CS relative to MATN stimulation or tone CSs. The results provide evidence that the auditory CS pathway necessary for delay eyeblink conditioning includes the MATN-LPN projection and the visual CS pathway includes the MPN.     

Simultaneous training on two hippocampus-dependent tasks facilitates acquisition of trace eyeblink conditioning

A common cellular alteration, reduced post-burst afterhyperpolarization (AHP) in CA1 neurons, is associated with acquisition of the hippocampus-dependent tasks trace eyeblink conditioning and the Morris water maze. As a similar increase in excitability is correlated with these two learning paradigms, we sought to determine the interactive behavioral effects of training animals on both tasks by using either a consecutive or simultaneous training design. In the consecutive design, animals were trained first on either the trace eyeblink conditioning task for six sessions, followed by training on the water maze task for six sessions, or vice versa. The simultaneous design consisted of six or 11 training days; animals received one session/day of both trace eyeblink conditioning and water maze training. Separate groups were used for consecutive and simultaneous training. Animals trained on both tasks simultaneously were significantly facilitated in their ability to acquire the trace eyeblink conditioning task; no effect of simultaneous training was seen on the water maze task. No effect was seen on acquisition for either task when using the consecutive training design. Taken together, these findings provide insight into how the hippocampus processes information when animals learn multiple hippocampus-dependent tasks.     

The effects of beta-noradrenergic receptor blockade on acquisition of eyeblink conditioning in 3-month-old F344 rats

There is evidence that blocking beta-noradrenergic receptors will cause deficits in some forms of learning. We investigated the effects of systemic injections of 1, 5, and 10 mg/kg doses of propranolol on acquisition of delay eyeblink conditioning in 3-month-old Fischer 344 rats. We presented a 3-kHz, 90-dB tone as a conditioning stimulus and a 6 psi airpuff as our unconditioned stimulus to freely moving rats. We monitored eyelid activity using EMG signals. The treatment subjects were injected with either propranolol or saline 0.5 h prior to daily training sessions. Two groups of control subjects, one receiving injections of saline and one receiving injections of 5 mg/kg propranolol, received daily training sessions with unpaired and randomized presentation of the tone and airpuff. Each daily training session for the treatment groups consisted of 27 paired training trials and 3 conditioned stimulus-alone training trials. Rats injected with saline vehicle or with 1 mg/kg propranolol achieved a 60% or better learned response rate within two training sessions. Rats injected with 5 or 10 mg/kg propranolol never achieved a response rate significantly different from animals that received unpaired, random presentations of the tone and airpuff stimuli. These results agree with prior studies from our lab that have shown a dose-dependent effect of beta-noradrenergic receptor blockade on learning in rabbit eyeblink conditioning as well as in a runway, motor learning paradigm. We believe that the beta-noradrenergic system plays an important role in learning and memory in more than one cerebellar-dependent learning paradigm.     

Lesions of the entorhinal cortex impair acquisition of hippocampal-dependent trace conditioning

Rabbits with the electrolytic lesions of bilateral entorhinal cortex (EC) were trained with the hippocampal-dependent trace conditioning of the nictitating membrane response. The multiple-unit activity of the hippocampal CA1 region was recorded during conditioning. The conditioned stimulus was a tone (1 kHz, 85 dB, 200-ms duration), the unconditioned stimulus was a corneal air puff (3 psi, 150-ms duration), and the interstimulus interval was 750 ms. The EC-lesioned animals showed only 30% conditioned response (CR) by the ninth session while the sham-operated animals showed above 80% CR. The lesioned animals did not show learning-related changes in the hippocampal activity. When the training was switched to the 300-ms interstimulus interval trace conditioning, both groups learned above 80% CR. The EC-lesioned animals, however, showed less learning-related activity in the hippocampus than the sham-operated animals. These results suggest that the development of the learning-related activity in the hippocampus depends on the intact EC, and that the EC may provide a possible pathway conveying learning information from the cerebellum or cerebral cortex to the hippocampus during the trace conditioning.     

d-cycloserine reverses the detrimental effects of stress on learning in females and enhances retention in males

Exposure to acute, inescapable stress produces a facilitation of subsequent classical eyeblink conditioning in male rats. The same stress exposure produces a profound deficit in classical eyeblink conditioning in females. Activation of N-methyl-d-aspartate receptors (NMDAr) is necessary for the effect of stress on learning in males while the contribution of NMDAr activation to the deficit in learning after stress is unknown. Here, we tested the influence of d-cycloserine (DCS), a positive modulator of the NMDAr, in stressed or unstressed male and female rats. Groups of males and females were exposed to an acute stressful event. One day later, they began training with four sessions of trace eyeblink conditioning. Each day before training, they were injected with DCS (15 mg/kg) or saline. Females treated with DCS during training responded similarly to those that were untreated. However, those that were stressed and the next day treated with the drug during training did not express the typical learning deficit, i.e. they learned to time the CR very well. Because the drug was administered well after the stressor, these data indicate that DCS reversed the negative effects of stress on learning in females. In males, the effect of DCS was subtle, resulting in higher asymptotic responding, and enhanced retention in a drug-free retention test. Thus, as shown previously, training in the presence of an NMDA receptor agonist enhances associative learning and memory retention. In addition, it can reverse learning deficits that have already been induced.     

Using hippocampal‐dependent eyeblink conditioning to predict individual differences in spatial reorientation strategies in 3‐ to 6‐year‐olds

The hippocampus is a subcortical structure in the medial temporal lobe involved in cognitive functions such as spatial navigation and reorientation, episodic memory, and associative learning. While much is understood about the role of hippocampal function in learning and memory in adults, less is known about the relations between the hippocampus and the development of these cognitive skills in young children due to the limitations of using standard methods (e.g., MRI) to examine brain structure and function in developing populations. This study used hippocampal‐dependent trace eyeblink conditioning (EBC) as a feasible approach to examine individual differences in hippocampal functioning as they relate to spatial reorientation and episodic memory performance in young children. Three‐ to six‐year‐old children (N = 50) completed tasks that measured EBC, spatial reorientation, and episodic memory, as well as non‐hippocampal‐dependent processing speed abilities. Results revealed that when age was held constant, individual differences in EBC performance were significantly related to individual differences in performance on the spatial reorientation test, but not on the episodic memory or processing speed tests. When the relations between hippocampal‐dependent EBC and different reorientation strategies were explored, it was found that individual differences in hippocampal function predicted the use of geometric information for reorienting in space as opposed to a combined strategy that uses both geometric information and salient visual cues. The utilization of eyeblink conditioning to examine hippocampal function in young populations and its implications for understanding the dissociation between spatial reorientation and episodic memory development are discussed.     

Hippocampal Theta Oscillations and Classical Conditioning

Studies are reviewed that support a hypothesized role for hippocampal theta oscillations in the neural plasticity underlying behavioral learning. Begun in Richard F. Thompson's laboratory in the 1970s, these experiments have documented a relationship between free-running 3- to 7-Hz hippocampal slow waves (theta) and rates of acquisition in rabbit classical nictitating membrane (NM) conditioning. Lesion and drug manipulations of septohippocampal projections have affected NM and jaw movement conditioning in ways consistent with a theta-related brain state being an important modulator of behavioral acquisition. These findings provide essential empirical support for the recently developed neurobiological and computational models that posit an important role for rhythmic oscillations (such as theta) in cellular plasticity and behavioral learning.     

Timing at the Start of Associative Learning

Some theories of associative learning imply that time plays a fundamental role in the acquisition process. Consistent with these theories, this paper presents evidence that the time from the onset of a conditioned stimulus (CS) until presentation of the unconditioned stimulus (US) is learned very rapidly at the start of training. We report two autoshaping studies and a study on aversive conditioning in goldfish in which we examine timing at the start of conditioning. We also review data from a number of other conditioning preparations, including fear-potentiated startle, appetitive conditioning in rats, and eyeblink conditioning in rabbits, that report conditioned response (CR) timing early in training. Acquisition speed and the very first expressions of conditioned responding often show sensitivity to the time of US presentation. In instances where temporal control is slowly expressed, it is likely due to performance factors, not to slow learning about time. In fact, the learning about time may be a necessary condition for associative learning.     

Effects of Early Hippocampal Lesions on Trace, Delay, and Long-Delay Eyeblink Conditioning in Developing Rats

The effects of bilateral hippocampal aspiration lesions on later acquisition of eyeblink conditioning were examined in developing Long-Evans rat pups. Lesions on postnatal day (PND) 10 were followed by evaluation of trace eyeblink conditioning (Experiment 1) and delay eyeblink conditioning (Experiment 2) on PND 25. Pairings of a tone conditioned stimulus (CS) and periocular shock unconditioned stimulus (US, 100 ms) were presented in one of three conditioning paradigms: trace (380 ms CS, 500 ms trace interval, 880 ms interstimulus interval [ISI]), standard delay (380 ms CS, 280 ms ISI), or long delay (980 ms CS, 880 ms ISI). The results of two experiments indicated that hippocampal lesions impaired trace eyeblink conditioning more than either type of delay conditioning. In light of our previous work on the ontogeny of trace, delay, and long-delay eyeblink conditioning (Ivkovich, Paczkowski, & Stanton, 2000) showing that trace and long-delay eyeblink conditioning had similar ontogenetic profiles, the current data suggest that during ontogeny hippocampal maturation may be more important for the short-term memory component than for the long-ISI component of trace eyeblink conditioning. The late development of conditioning over long ISIs may depend on a separate process such as protracted development of cerebellar cortex.     

Ontogenetic changes in the neural mechanisms of eyeblink conditioning

The rodent eyeblink conditioning paradigm is an ideal model system for examining the relationship between neural maturation and the ontogeny of associative learning. Elucidation of the neural mechanisms underlying the ontogeny of learning is tractable using eyeblink conditioning because the necessary neural circuitry (cerebellum and interconnected brainstem nuclei) underlying the acquisition and retention of the conditioned response (CR) has been identified in adult organisms. Moreover, the cerebellum exhibits substantial postnatal anatomical and physiological maturation in rats. The eyeblink CR emerges developmentally between postnatal day (PND) 17 and 24 in rats. A series of experiments found that the ontogenetic emergence of eyeblink conditioning is related to the development of associative learning and not related to changes in performance. More recent studies have examined the relationship between the development of eyeblink conditioning and the physiological maturation of the cerebellum, a brain structure that is necessary for eyeblink conditioning in adult organisms. Disrupting cerebellar development with lesions or antimitotic treatments impairs the ontogeny of eyeblink conditioning. Studies of the development of physiological processes within the cerebellum have revealed striking ontogenetic changes in stimulus-elicited and learning-related neuronal activity. Neurons in the interpositus nucleus and Purkinje cells in the cortex exhibit developmental increases in neuronal discharges following the unconditioned stimulus (US) and in neuronal discharges that model the amplitude and time-course of the eyeblink CR. The developmental changes in CR-related neuronal activity in the cerebellum suggest that the ontogeny of eyeblink conditioning depends on the development of mechanisms that estavlish cerebellar plasticity. Learning and the induction of neural plasticity depend on the magnitude of the US input to the cerebellum. The role of developmental changes in the efficacy of the US pathway has been investigated by monitoring neuronal activity in the inferior olive and with stimulation techniques. The results of these experiments indicate that the development of the conditioned eyeblink response may depend on dynamic interactions between multiple developmental processes within the eyeblink neural circuitry.     

Classical Delay Eyeblink Conditioning in in 4- and 5-Month-Old Human Infants

Abstract-Simple delay classical eyeblink conditioning, using a tone conditioned stimulus (CS) and airpuff unconditioned stimulus (US), was studied in cross-sectional samples of 4- and 5-month-old healthy, full-term infants. Infants received two identical training sessions, 1 week apart. At both ages, infants experiencing paired tones and air-puffs demonstrated successful conditioning over two sessions, relative to control subjects who had unpaired training. Conditioning was not evident, however, during the first session. Two additional groups of 5-month-olds received varied experiences during Session 1, either unpaired presentations of the CS and US or no stimulus exposure, fol-lowed by paired conditioning during Session 2. Results from these groups suggest that the higher level of conditioning observed following two sessions of paired conditioning was not the result of familiarity with the testing environment or the stimuli involved but, rather, the result of retention of associative learning not expressed during the first conditioning session.     

Changing the rate and hippocampal dependence of trace eyeblink conditioning: slow learning enhances survival of new neurons

Trace eyeblink conditioning in which a conditioned stimulus and unconditioned stimulus are separated by a gap, is hippocampal dependent and can rescue new neurons in the adult dentate gyrus from death (e.g., Beylin et al., 2001; Gould et al., 1999). Tasks requiring more training trials for reliable expression of the conditioned response are most effective in enhancing survival of neurons (Waddell & Shors, 2008). To dissociate hippocampal dependence from acquisition rate, we facilitated hippocampal-dependent trace eyeblink conditioning in two ways: a shorter trace interval and signaling the intertrial interval with a post-US cue. Trace conditioning with a shorter trace interval (250ms) requires an intact hippocampus, and acquisition is faster relative to rats trained with a 500ms trace interval (e.g., Weiss et al., 1999). Using excitotoxic hippocampal lesions, we confirmed that eyeblink conditioning with the 250 or 500ms trace interval is hippocampal dependent. However, training with the post-US cue was not hippocampal dependent. The majority of lesion rats in this condition reached criterion of conditioned responding. To determine whether hippocampal dependence is sufficient to rescue adult-generated neurons in the dentate gyrus, rats were injected with BrdU and trained in one of the three trace eyeblink arrangements one week later. Of these training procedures, only the 500ms trace interval enhanced survival of new cells; acquisition of this task proceeded slowly relative to the 250ms and post-US cue conditions. These data demonstrate that rate of acquisition and not hippocampal dependence determines the impact of learning on adult neurogenesis.     

Ontogenetic Changes in the Neural Mechanisms of Eyeblink Conditioning

The rodent eyeblink conditioning paradigm is an ideal model system for examining the relationship between neural maturation and the ontogeny of associative learning. Elucidation of the neural mechanisms underlying the ontogeny of learning is tractable using eyeblink conditioning because the necessary neural circuitry (cerebellum and interconnected brainstem nuclei) underlying the acquisition and retention of the conditioned response (CR) has been identified in adult organisms. Moreover, the cerebellum exhibits substantial postnatal anatomical and physiological maturation in rats. The eyeblink CR emerges developmentally between postnatal day (PND) 17 and 24 in rats. A series of experiments found that the ontogenetic emergence of eyeblink conditioning is related to the development of associative learning and not related to changes in performance. More recent studies have examined the relationship between the development of eyeblink conditioning and the physiological maturation of the cerebellum, a brain structure that is necessary for eyeblink conditioning in adult organisms. Disrupting cerebellar development with lesions or antimitotic treatments impairs the ontogeny of eyeblink conditioning. Studies of the development of physiological processes within the cerebellum have revealed striking ontogenetic changes in stimulus-elicited and learning-related neuronal activity. Neurons in the interpositus nucleus and Purkinje cells in the cortex exhibit developmental increases in neuronal discharges following the unconditioned stimulus (US) and in neuronal discharges that model the amplitude and time-course of the eyeblink CR. The developmental changes in CR-related neuronal activity in the cerebellum suggest that the ontogeny of eyeblink conditioning depends on the development of mechanisms that establish cerebellar plasticity. Learning and the induction of neural plasticity depend on the magnitude of the US input to the cerebellum. The role of developmental changes in the efficacy of the US pathway has been investigated by monitoring neuronal activity in the inferior olive and with stimulation techniques. The results of these experiments indicate that the development of the conditioned eyeblink response may depend on dynamic interactions between multiple developmental processes within the eyeblink neural circuitry.     

Average group behavior does not represent individual behavior in classical conditioning of the honeybee

Conditioned behavior as observed during classical conditioning in a group of identically treated animals provides insights into the physiological process of learning and memory formation. However, several studies in vertebrates found a remarkable difference between the group-average behavioral performance and the behavioral characteristics of individual animals. Here, we analyzed a large number of data (1640 animals) on olfactory conditioning in the honeybee (Apis mellifera). The data acquired during absolute and differential classical conditioning differed with respect to the number of conditioning trials, the conditioned odors, the intertrial intervals, and the time of retention tests. We further investigated data in which animals were tested for spontaneous recovery from extinction. In all data sets we found that the gradually increasing group-average learning curve did not adequately represent the behavior of individual animals. Individual behavior was characterized by a rapid and stable acquisition of the conditioned response (CR), as well as by a rapid and stable cessation of the CR following unrewarded stimuli. In addition, we present and evaluate different model hypotheses on how honeybees form associations during classical conditioning by implementing a gradual learning process on the one hand and an all-or-none learning process on the other hand. In summary, our findings advise that individual behavior should be recognized as a meaningful predictor for the internal state of a honeybee--irrespective of the group-average behavioral performance.     

Trace eyeblink conditioning requires the hippocampus but not autophosphorylation of alphaCaMKII in mice

Little is known about signaling mechanisms underlying temporal associative learning. Here, we show that mice with a targeted point mutation that prevents autophosphorylation of alphaCaMKII (alphaCaMKII(T286A)) learn trace eyeblink conditioning normally. This forms a sharp contrast to the severely impaired spatial learning in the water maze and contextual fear conditioning observed in alphaCaMKII(T286A) mutants. Importantly, hippocampal lesions impaired trace eyeblink conditioning in alphaCaMKII(T286A) mice, suggesting a potential role of hippocampal alphaCaMKII-independent mechanisms. These results indicate that hippocampal signaling mechanisms that underlie temporal associative learning as assessed by trace eyeblink conditioning may differ from those of spatial and contextual learning.     

Galantamine facilitates acquisition of a trace-conditioned eyeblink response in healthy, young rabbits

Previous work has demonstrated that drugs increasing brain concentrations of acetylcholine can enhance cognition in aging and brain-damaged organisms. The present study assessed whether galantamine (GAL), an allosteric modulator of nicotinic cholinergic receptors and weak acetylcholinesterase inhibitor, could improve acquisition and retention of an eyeblink (EB) classical conditioning task in healthy, young animals. We trained 24 rabbits (n = 8/group) in a 1000-msec trace Pavlovian EB conditioning paradigm in which a tone conditioned stimulus (CS) was presented for 500 msec, followed by a 500-msec trace period in which no stimuli were presented. A 100-msec corneal airpuff was the unconditioned stimulus (US). Acquisition sessions, consisting of 100 trials each, occurred daily for 10 consecutive days, followed by 3 d of extinction training. Animals were treated with one of three doses of GAL (0.0-3.0 mg/kg) prior to each session. Animals that received 3.0 mg/kg GAL showed significantly more EB conditioned responses (CRs) in fewer training trials than animals receiving either 1.5 mg/kg GAL or vehicle injections. GAL had no effect on CR performance during extinction. Pseudoconditioning control experiments, consisting of 200 explicitly unpaired tone-puff presentations indicated that GAL did not increase reactivity to the CS or US. These findings indicate that GAL may improve acquisition of moderately difficult associative learning tasks in healthy young organisms.