Matthew M. Campolattaro

Eyeblink conditioning in 12-day-old rats using pontine stimulation as the conditioned stimulus

A fundamental issue in developmental science is whether ontogenetic changes in memory are caused by the development of cellular plasticity mechanisms within the brains memory systems or maturation of sensory inputs to the memory systems. Here, we provide evidence that the development of eyeblink conditioning, a form of associative learning that depends on the cerebellum, is driven by the development of sensory inputs rather than the development of neuronal plasticity mechanisms. We find that rats as young as 12 days old show associative eyeblink conditioning when pontine stimulation is used in place of an external (e.g., a tone) conditioned stimulus. Eyeblink-conditioned responses established with pontine stimulation in 12-day-old rats were reversibly abolished by an infusion of muscimol into the cerebellar interpositus nucleus. The findings suggest that cerebellar neurons are capable of supporting associative learning-specific plasticity in vivo in very immature animals if given sufficient afferent stimulation.

Neuronal Correlates of Cross-Modal Transfer in the Cerebellum and Pontine Nuclei

Cross-modal transfer occurs when learning established with a stimulus from one sensory modality facilitates subsequent learning with a new stimulus from a different sensory modality. The current study examined neuronal correlates of cross-modal transfer of pavlovian eyeblink conditioning in rats. Neuronal activity was recorded from tetrodes within the anterior interpositus nucleus (IPN) of the cerebellum and basilar pontine nucleus (PN) during different phases of training. After stimulus preexposure and unpaired training sessions with a tone conditioned stimulus (CS), light CS, and periorbital stimulation unconditioned stimulus (US), rats received associative training with one of the CSs and the US (CS1–US). Training then continued on the same day with the other CS to assess cross-modal transfer (CS2–US). The final training session included associative training with both CSs on separate trials to establish stronger cross-modal transfer (CS1/CS2). Neurons in the IPN and PN showed primarily unimodal responses during pretraining sessions. Learning-related facilitation of activity correlated with the conditioned response (CR) developed in the IPN and PN during CS1–US training. Subsequent CS2–US training resulted in acquisition of CRs and learning-related neuronal activity in the IPN but substantially less little learning-related activity in the PN. Additional CS1/CS2 training increased CRs and learning-related activity in the IPN and PN during CS2–US trials. The findings suggest that cross-modal neuronal plasticity in the PN is driven by excitatory feedback from the IPN to the PN. Interacting plasticity mechanisms in the IPN and PN may underlie behavioral cross-modal transfer in eyeblink conditioning.

Perirhinal cortex lesions impair feature-negative discrimination

The role of the perirhinal cortex in inhibitory eyeblink conditioning was examined. In Experiment 1, rats were given lesions of the perirhinal cortex or control surgery and subsequently trained with a feature-negative discrimination procedure followed by summation and retardation tests for conditioned inhibition. Perirhinal cortex lesions impaired, but did not prevent acquisition of feature-negative discrimination. Results from the summation test showed that rats with perirhinal cortex lesions could not generalize feature-negative discrimination to a new stimulus. There were no group differences during the retardation test. Experiment 2 showed that lesions of the perirhinal cortex did not impair simple excitatory conditioning. Experiment 3 showed that perirhinal cortex lesions had no effect on acquisition of a simple tone-light discrimination. The results suggest that the perirhinal cortex plays a role in eyeblink conditioning when using discrimination procedures involving overlapping stimuli.

Cerebellar inactivation impairs cross modal savings of eyeblink conditioning.

Eyeblink conditioning using a conditioned stimulus (CS) from one sensory modality (e.g., an auditory CS) is greatly enhanced when the subject is previously trained with a CS from a different sensory modality (e.g., a visual CS). The enhanced acquisition to the second modality CS results from cross modal savings. The current study was designed to examine the role of the cerebellum in establishing cross modal savings in eyeblink conditioning with rats. In the first experiment rats were given paired or unpaired presentations with a CS (tone or light) and an unconditioned stimulus. All rats were then given paired training with a different modality CS. Only rats given paired training showed cross modal savings to the second modality CS. Experiment 2 showed that cerebellar inactivation during initial acquisition to the first modality CS completely prevented savings when training was switched to the second modality CS. Experiment 3 showed that cerebellar inactivation during initial cross modal training also prevented savings to the second modality stimulus. These results indicate that the cerebellum plays an essential role in establishing cross modal savings of eyeblink conditioning.

Perirhinal cortex lesions impair simultaneous but not serial feature-positive discrimination learning.

The role of the perirhinal cortex in discriminative eyeblink conditioning was examined by means of feature-positive discrimination procedures with simultaneous (A-/XA+) and serial (A-/X-->A+) stimulus compounds. Lesions of the perirhinal cortex severely impaired acquisition of simultaneous feature-positive discrimination but produced no impairment in serial feature-positive discrimination. The results suggest that the perirhinal cortex plays a role in discriminative eyeblink conditioning by resolving ambiguity in discriminations with overlapping stimulus elements.

Ontogenetic change in the auditory conditioned stimulus pathway for eyeblink conditioning

Two experiments examined the neural mechanisms underlying the ontogenetic emergence of auditory eyeblink conditioning. Previous studies found that the medial auditory thalamus is necessary for eyeblink conditioning with an auditory conditioned stimulus (CS) in adult rats. In experiment 1, stimulation of the medial auditory thalamus was used as a CS in rat pups trained on postnatal days (P) 17-18, 24-25, or 31-32. All three age groups showed significant acquisition relative to unpaired controls. However, there was an age-related increase in the rate of conditioning. Experiment 2 examined the effect of inactivating the medial auditory thalamus with muscimol on auditory eyeblink conditioning in rats trained on P17-18, 24-25, or 31-32. Rat pups trained on P24-25 and P31-32, but not P17-18, showed a significant reduction in conditioned responses following muscimol infusions. The findings suggest that the thalamic contribution to auditory eyeblink conditioning continues to develop through the first postnatal month.

Changes in inhibition during differential eyeblink conditioning with increased training

Three experiments examined inhibitory learning in rats, using Pavlovian and differential inhibitory eyeblink conditioning procedures. Experiment 1 was designed to compare summation and retardation effects following Pavlovian conditioned inhibition (A1/XA) or differential inhibition (A1/X) training using auditory and visual conditioned stimuli (CSs). After ten 100-trial sessions of training, both Pavlovian conditioned inhibition and differential inhibition produced a retardation effect. However, a summation effect was obtained only for rats given Pavlovian conditioned inhibition training. Experiment 2 showed that increasing differential inhibition training to twenty 100-trial sessions produced summation and retardation effects. In Experiment 3, rats were trained with either ten or twenty 100-trial sessions of intramodal inhibitory training with two tone CSs (2 kHz vs. 8 kHz). Summation and retardation effects were obtained after only 20 sessions of differential conditioning. The findings indicate that extensive training is needed to establish conditioned inhibition with intermodal or intramodal differential conditioning.

Examination of bilateral eyeblink conditioning in rats.

This experiment monitored eyelid responses bilaterally during delay eyeblink conditioning in rats. Rats were given paired or unpaired training with a tone or light conditioned stimulus (CS) and a unilateral periorbital shock unconditioned stimulus (US). Rats given paired training acquired high levels of conditioned responses (CRs), which occurred in both eyelids. However, acquisition was faster, and the overall percentage of CRs was greater in the eyelid that was ipsilateral to the US. CRs in the eyelid ipsilateral to the US also had shorter onset latencies and larger amplitudes than CRs in the contralateral eyelid. Both eyelids consistently showed high percentages of unconditioned responses (UR) to the US, and the UR amplitude decreased across training sessions in the paired group. The present study demonstrated that CRs occur robustly in both eyelids of rats given eyeblink conditioning, which is similar to previous findings in humans and monkeys. The results also showed that conditioning occurs more prominently in the eyelid that is ipsilateral to the US, which is similar to previous findings in humans, monkeys, dogs, and rabbits.

Learning-related neuronal activity in the ventral lateral geniculate nucleus during associative cerebellar learning.

During delay eyeblink conditioning, rats learn to produce an eyelid-closure conditioned response (CR) to a conditioned stimulus (CS), such as a light, which precedes and coterminates with an unconditioned stimulus (US). Previous studies have suggested that the ventral lateral geniculate nucleus (LGNv) might play an important role in visual eyeblink conditioning by supplying visual sensory input to the pontine nuclei (PN) and also receiving feedback from the cerebellum. No prior study has investigated LGNv neuronal activity during eyeblink conditioning. The present study used multiple tetrodes to monitor single-unit activity in the rat LGNv during pre-exposure (CS only), unpaired CS/US, and paired CS-US training conditions. This behavioral-training sequence was used to investigate nonassociative- and associative-driven neuronal activity in the LGNv during training. LGNv neuronal activity habituated during unpaired training and then recovered from habituation during subsequent paired training, which may indicate that the LGNv plays a role in attention to the CS. The amplitude of LGNv neuronal activity correlated with CR production during paired but not unpaired CS/US training. Cerebellar feedback to the LGNv may play a role in modulating LGNv activity and attention to the CS during paired training. Based on the present findings, we hypothesize that the role of LGNv in visual eyeblink conditioning goes beyond simply routing visual CS information to the PN and involves modulation of attention.

Sex differences and the role of acute stress in the open-field tower maze

Many studies provide evidence that differences in spatial learning exist between males and females. However, it is necessary to consider non-mnemonic factors that may influence these findings. The present experiment investigated acquisition, retention, and the effects of stress on response- and place-learning in male and female rats. Rats were trained in an open-field tower maze. Procedures were used to minimize stress in the rats, and their ability to solve place- or response-learning in the maze was determined by analyzing a response variable (i.e., first choice correct response) that was not influenced by general locomotor activity. The results revealed that male and female rats acquire place- and response-learning at the same rate even though females moved significantly faster in the maze. However, females showed better retrieval of place-, but not response-learning compared to male rats. This effect appeared to be enhanced when the rats were tested immediately following an acute restraint stress. Furthermore, both female and male rats that were exposed to acute restraint stress showed less impairment than controls when subsequently tested in a novel situation. These findings have clinical implications that a mild physiological stress response can make one more cognitively resistant to adversities later in life.