Percentage Of Conditioned Responses Research Articles

Neurite density of the hippocampus is associated with trace eyeblink conditioning latency in 4- to 6-year-olds.

Limited options exist to evaluate the development of hippocampal function in young children. Research has established that trace eyeblink conditioning (EBC) relies on a functional hippocampus. Hence, we set out to investigate whether trace EBC is linked to hippocampal structure, potentially serving as a valuable indicator of hippocampal development. Our study explored potential associations between individual differences in hippocampal volume and neurite density with trace EBC performance in young children. We used onset latency of conditioned responses (CR) and percentage of conditioned responses (% CR) as measures of hippocampal-dependent associative learning. Using a sample of typically developing children aged 4 to 6years (N = 30; 14 girls; M = 5.70years), participants underwent T1- and diffusion-weighted MRI scans and completed a 15-min trace eyeblink conditioning task conducted outside the MRI. % CR and CR onset latency were calculated based on all trials involving tone-puff presentations and tone-alone trials. Findings revealed a connection between greater left hippocampal neurite density and delayed CR onset latency. Children with higher neurite density in the left hippocampus tended to blink closer to the onset of the unconditioned stimulus, indicating that structural variations in the hippocampus were associated with more precise timing of conditioned responses. No other relationships were observed between hippocampal volume, cerebellum volume or neurite density, hippocampal white matter connectivity and any EBC measures. Preliminary results suggest that trace EBC may serve as a straightforward yet innovative approach for studying hippocampal development in young children and populations with atypical development.

Sleep deprivation directly following eyeblink-conditioning impairs memory consolidation.

The relation between sleep and different forms of memory formation continues to be a relevant topic in our daily life. Sleep has been found to affect cerebellum-dependent procedural memory formation, but it remains to be elucidated to what extent the level of sleep deprivation directly after motor training also influences our ability to store and retrieve memories. Here, we studied the effect of disturbed sleep in mice during two different time-windows, one covering the first four hours following eyeblink conditioning (EBC) and another window following the next period of four hours. Compared to control mice with sleep ad libitum, the percentage of conditioned responses and their amplitude were impaired when mice were deprived of sleep directly after conditioning. This impairment was still significant when the learned EBC responses were extinguished and later reacquired. However, consolidation of eyeblink responses was not affected when mice were deprived later than four hours after acquisition, not even when tested during a different day-night cycle for control. Moreover, mice that slept longer directly following EBC showed a tendency for more conditioned responses. Our data indicate that consolidation of motor memories can benefit from sleep directly following memory formation.

Impaired cerebellar plasticity and eye-blink conditioning in calpain-1 knock-out mice.

Calpain-1 and calpain-2 are involved in the regulation of several signaling pathways and neuronal functions in the brain. Our recent studies indicate that calpain-1 is required for hippocampal synaptic plasticity, including long-term depression (LTD) and long-term potentiation (LTP) in field CA1. However, little is known regarding the contributions of calpain-1 to cerebellar synaptic plasticity. Low frequency stimulation (LFS, 5 Hz, 5 min)-induced LTP at parallel fibers to Purkinje cell synapses was markedly impaired in cerebellar slices from calpain-1 knock-out (KO) mice. Application of a selective calpain-2 inhibitor enhanced LFS-induced LTP in both wild-type (WT) and calpain-1 KO mice. Three protocols were used to induce LTD at these synapses: LFS (1 Hz, 15 min), perfusion with high potassium and glutamate (K-Glu) or dihydroxyphenylglycine (DHPG), a mGluR1 agonist. All three forms of LTD were impaired in calpain-1 KO mice. DHPG application stimulated calpain-1 but not calpain-2 in cerebellar slices, and DHPG-induced LTD impairment was reversed by application of a protein phosphatase 2A (PP2A) inhibitor, okadaic acid. As in hippocampus, BDNF induced calpain-1 activation and PH domain and Leucine-rich repeat Protein Phosphatase 1/suprachiasmatic nucleus oscillatory protein (PHLPP1/SCOP) degradation followed by extracellular signal-regulated kinase (ERK) activation, as well as calpain-2 activation leading to degradation of phosphatase and tensin homolog deleted on chromosome ten (PTEN) in cerebellar slices. The role of calpain-1 in associative learning was evaluated in the delay eyeblink conditioning (EBC). Calpain-1 KO mice exhibited significant learning impairment in EBC during the first 2 days of acquisition training. However, after 5 days of training, the percentage of conditioned responses (CRs) between calpain-1 KO and WT mice was identical. Both calpain-1 KO and WT mice exhibited typical extinction patterns. Our results indicate that calpain-1 plays critical roles in multiple forms of synaptic plasticity and associative learning in both hippocampus and cerebellum.

Awareness is essential for differential delay eyeblink conditioning with soft-tone but not loud-tone conditioned stimuli.

The role of awareness in differential delay eyeblink conditioning (DEC) remains controversial. Here, we investigated the involvement of awareness in differential DEC with a soft or a loud tone as the conditioned stimulus (CS). In the experiment, 36 participants were trained in differential DEC with a soft tone (60 dB) or a loud tone (85 dB) as the CS, paired with a corneal air-puff as the unconditioned stimulus (US). After conditioning, awareness of the relationship between the CS and the US was assessed with a 17-item true/false questionnaire. Interestingly, during differential DEC with a soft-tone CS, a higher proportion of differential conditioned responses (CRs) was evident in participants who were aware than those who were unaware. In contrast, when a loud tone was used as the CS, the proportion of differential CRs of the aware participants did not differ significantly from those who were unaware over any of the blocks of 20 trials. In unaware participants, the percentage of differential CRs with a loud-tone CS was significantly higher than that with a soft-tone CS; however in participants classified as aware, the percentage of differential CRs with a loud-tone CS did not differ significantly from that with a soft-tone CS. The present findings suggest that awareness is critical for differential DEC when the delay task is rendered more difficult.

The Rostral Medial Prefrontal Cortex Regulates the Expression of Conditioned Eyelid Responses in Behaving Rabbits

We studied the contribution of the rostral mPFC (rmPFC) to the acquisition and performance of classical eyeblink conditioning in rabbits using a delay paradigm. The rmPFC was determined by its afferent projections from the medial half of the mediodorsal thalamic nucleus. The rmPFC neurons were identified by their antidromic activation from the mediodorsal nucleus and/or by their firing characteristics. The rmPFC neurons increased their firing during the first conditioning sessions, but decreased it when conditioned responses (CRs) reached asymptotic values. Therefore, no significant relationships could be established between neuronal firing rates and the percentage of CRs or the electromyographic (EMG) activity of the orbicularis oculi muscle during conditioning. Electrical train stimulation of the rmPFC produced a significant inhibition of air-puff-evoked blinks and reduced the generation of CRs compared with controls. Inhibition of the rmPFC by the local injection of lidocaine produced an increase in the amplitude of evoked reflex and conditioned eyeblinks and in the percentage of CRs. The rmPFC seems to be a potent inhibitor of reflex and conditioned eyeblinks, controlling the release of newly acquired eyelid responses until advanced stages of the acquisition process--i.e., until the need for the acquired response is fully confirmed. Therefore, the rmPFC seems to act as a "flip-flop" mechanism in controlling behavior.

Role of Reuniens Nucleus Projections to the Medial Prefrontal Cortex and to the Hippocampal Pyramidal CA1 Area in Associative Learning

We studied the interactions between short- and long-term plastic changes taking place during the acquisition of a classical eyeblink conditioning and following high-frequency stimulation (HFS) of the reuniens nucleus in behaving mice. Synaptic changes in strength were studied at the reuniens-medial prefrontal cortex (mPFC) and the reuniens-CA1 synapses. Input/output curves and a paired-pulse study enabled determining the functional capabilities of the two synapses and the optimal intensities to be applied at the reuniens nucleus during classical eyeblink conditioning and for HFS applied to the reuniens nucleus. Animals were conditioned using a trace paradigm, with a tone as conditioned stimulus (CS) and an electric shock to the trigeminal nerve as unconditioned stimulus (US). A single pulse was presented to the reuniens nucleus to evoke field EPSPs (fEPSPs) in mPFC and CA1 areas during the CS-US interval. No significant changes in synaptic strength were observed at the reuniens-mPFC and reuniens-CA1 synapses during the acquisition of eyelid conditioned responses (CRs). Two successive HFS sessions carried out during the first two conditioning days decreased the percentage of CRs, without evoking any long-term potentiation (LTP) at the recording sites. HFS of the reuniens nucleus also prevented the proper acquisition of an object discrimination task. A subsequent study revealed that HFS of the reuniens nucleus evoked a significant decrease of paired-pulse facilitation. In conclusion, reuniens nucleus projections to prefrontal and hippocampal circuits seem to participate in the acquisition of associative learning through a mechanism that does not required the development of LTP.

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.

Exceptional good cognitive and phenotypic profile in a male carrying a mosaic mutation in the FMR1 gene

Fragile X (FRAX) syndrome is a commonly inherited form of mental retardation resulting from the lack of expression of the fragile X mental retardation protein (FMRP). It is caused by a stretch of CGG repeats within the fragile X gene, which can be unstable in length as it is transmitted from generation to generation. Once the repeat exceeds a threshold length, the FMR1 gene is methylated and no protein is produced resulting in the fragile X phenotype. The consequences of FMRP absence in the mechanisms underlying mental retardation are unknown. We have identified a male patient in a classical FRAX family without the characteristic FRAX phenotype. His intelligence quotient (IQ) is borderline normal despite the presence of a mosaic pattern of a pre-mutation (25%), full mutation (60%) and a deletion (15%) in the FMR1 gene. The cognitive performance was determined at the age of 28 by the Raven test and his IQ was 81. However, FMRP expression studies in both hair roots and lymphocytes, determined at the same time as the IQ test, were within the affected male range. The percentage of conditioned responses after delay eyeblink conditioning was much higher than the average percentage measured in FRAX studies. Moreover, this patient showed no correlation between FMRP expression and phenotype and no correlation between DNA diagnostics and phenotype.

Deletion of FMR1 in Purkinje Cells Enhances Parallel Fiber LTD, Enlarges Spines, and Attenuates Cerebellar Eyelid Conditioning in Fragile X Syndrome

Absence of functional FMRP causes Fragile X syndrome. Abnormalities in synaptic processes in the cerebral cortex and hippocampus contribute to cognitive deficits in Fragile X patients. So far, the potential roles of cerebellar deficits have not been investigated. Here, we demonstrate that both global and Purkinje cell-specific knockouts of Fmr1 show deficits in classical delay eye-blink conditioning in that the percentage of conditioned responses as well as their peak amplitude and peak velocity are reduced. Purkinje cells of these mice show elongated spines and enhanced LTD induction at the parallel fiber synapses that innervate these spines. Moreover, Fragile X patients display the same cerebellar deficits in eye-blink conditioning as the mutant mice. These data indicate that a lack of FMRP leads to cerebellar deficits at both the cellular and behavioral levels and raise the possibility that cerebellar dysfunctions can contribute to motor learning deficits in Fragile X patients.

Performance of α7 nicotinic receptor null mutants is impaired in appetitive learning measured in a signaled nose poke task

Wild-type and mutant mice lacking expression of α5, α7, β2, β3, or β4 neuronal nicotinic cholinergic receptors (nAChRs) were compared on a signaled nose poke task, a multi-phased task used to measure appetitive learning and impulsivity. In the early phases of training, mutants of all nicotinic lines did not differ compared to wild types in the days to reach criterion when mice were required to nose poke for a sucrose reward on FR1 or FR3 schedules, or in their ability to respond to an auditory clicker to receive a sucrose reward. However, mutants lacking α7 nAChRs, but not lines lacking other nAChRs, showed impairments when task difficulty was increased such that an auditory stimulus was presented on a variable schedule and mice were required to withhold their responses until the presentation of the auditory cue to obtain a reward. α7 mutants were impaired compared to wild types in appetitive learning as measured by the percentage of conditioned responses but overcame their deficits with extensive training for 10 days. However, when efficiency ratios were used to measure impulsivity, α7 mutants exhibited lower efficiency ratios even after 10 days of training. These results support a role of the α7 nicotinic receptor in mediating appetitive learning and suggest a potential role for the α7 nAChRs in the regulation of behavioral disinhibition.

Effects of a Corneal Anesthetic on Extinction of the Classically Conditioned Nictitating Membrane Response in the Rabbit (Oryctolagus cuniculus).

Rabbits (Oryctolagus cuniculus) were presented with 7 daily sessions of tone-alone training after conditioning. Before the beginning of each of the first 4 extinction sessions, an artificial tear solution or tetracaine hydrochloride was administered to the cornea of rabbits in the control group (n = 6) and experimental group (n = 7), respectively. There were no between-group differences in the percentage of conditioned responses between both groups. However, the amplitude of the conditioned response was notably reduced in the tetracaine group (M = 0.40, SEM +/- 0.216) relative to the control group (M = 1.32, SEM +/- 0.639) early in extinction. Results seem to suggest that although motor output has been found to play an important role in extinction, corneal sensory feedback is not necessary.

MRI-Assessed Volume of Cerebellum Correlates with Associative Learning

Richard F. Thompson's cerebellar model of classical eyeblink conditioning highlights Purkinje cells in cerebellar cortex and principal cells in the deep cerebellar nucleus as the integrating cells for acquisition of conditioned responses (CRs). CR acquisition is significantly slower in rabbits with lesions to cerebellar cortex and in Purkinje cell-deficient mice that lose all cerebellar cortical Purkinje cells. Purkinje cells are the largest neurons in the cerebellum and contribute significantly to cerebellar volume. Magnetic resonance imaging (MRI) was used to assess cerebellar volume in humans. Cerebellar volume was related to eyeblink conditioning (400-ms delay procedure) in 8 adults (21–35 years) and compared to 8 older adults (77–95 years) tested previously (Woodruff-Pak, Goldenberg, Downey-Lamb, Boyko, & Lemieux, 2000). In the young adult sample, there was a high correlation between percentage of CRs in a session and cerebellar volume (corrected for total intracranial volume [TIV], r = .58, p = .066). There were statistically significant age differences in cerebellar volume, t(14) = 8.96, p < .001, and percentage of CRs, t(14) = 3.85, p < .002, but no age difference in TIV. Combining the young and older adult sample, the correlation between percentage of CRs and cerebellar volume (corrected for TIV) was .832 (p < .001). Cerebellar volume showed age-related deficits likely due to Purkinje cell loss. Individual differences in classical eyeblink conditioning are associated with differences in cerebellar volume, supporting Thompson's model of a cerebellar cortical role in facilitating this form of associative learning.

Nicotinic modulation in an animal model of a form of associative learning impaired in Alzheimer’s disease

Eyeblink classical conditioning is a widely used associative learning paradigm that has striking behavioral and neurobiological parallels between humans and other mammals. Eyeblink conditioning is impaired in older organisms, and patients with Alzheimer’s disease (AD) are impaired beyond the normal aging deficit. The cholinergic system is of demonstrated involvement in eyeblink conditioning. Blockade of nicotinic cholinergic receptors with mecamylamine prolonged acquisition of conditioned responses (CRs) in young adult rabbits, and the nicotinic agonist, GTS-21 ameliorated conditioning deficits in older rabbits. Galantamine induces allosteric modulation of nicotinic cholinergic receptors to increase acetylcholine release as well as acting as an acetylcholinesterase inhibitor. Galantamine doses of 0.0, 1.0, 2.0, 3.0, and 4.0 mg/kg were tested in ten daily sessions in 40 retired breeder rabbits (mean age=29 months) in the 750 ms delay conditioning paradigm. A dose of 3 mg/kg galantamine was effective in improving conditioning in older rabbits, enabling them to achieve learning criterion rapidly and to produce a very high percentage of CRs. Control tests of rabbits in explicitly unpaired conditions demonstrated that non-associative factors could not account for the results. The efficacy of galantamine in a learning paradigm that shows severe impairment in AD indicates that the drug may be effective as a cognition-enhancer in AD.

Increase of extracellular dopamine in prefrontal cortex of gerbils during acquisition of the avoidance strategy in the shuttle-box

We examined whether extracellular dopamine (DA) increase in medial prefrontal cortex is correlated with the establishment or with the retrieval of an auditory avoidance strategy in a shuttle-box. Using microdialysis from right medial prefrontal cortex (mPFC) in combination with behavioral measures, gerbils were trained on a tone-footshock combination in two sessions on two successive days. Sessions in each animal involving avoidance strategy formation and avoidance strategy retrieval were differentiated post hoc by using the percentage of conditioned responses (CR) and other behavioral measures of training sessions. It was found that exclusively strategy formation correlated with high DA levels in mPFC.

Hippocampal Acetylcholine Increases During Eyeblink Conditioning in the Rabbit

The classically conditioned rabbit nictitating membrane reflex (NMR) is modulated by the septohippocampal cholinergic system. Disruption of this system retards NMR acquisition. Aluminum (Al) is a neurotoxin that interferes with hippocampal acetylcholine (ACh) synthesis and release. Using microdialysis, this study tested the hypothesis that NMR acquisition in the rabbit is associated with hippocampal ACh release. This was conducted by measuring ACh release in control and Al-intoxicated rabbits during NMR training. NMR training consisted of four sessions of 100 conditioning trials/session in a delay paradigm. The percentage of conditioned responses (CRs) increased with each conditioning session campus increased significantly over baseline in the control group during the second and third conditioning sessions. In the Alintoxicated group, ACh release did not increase significantly during any conditioning session. A separate group of rabbits was pseudoconditioned, receiving the same conditioning stimuli, although explicitly unpaired. This group did not acquire the CR. Acetylcholine release did not significantly increase during any conditioning session, suggesting that the increase in ACh release observed in the control group was not merely a product of conditioning stimuli presentation. The lack of increased ACh release in the Al-intoxicated rabbits was associated with a CR acquisition deficit. The results of this study are consistent with a role of hippocampal cholinergic function in NMR acquisition in the rabbit.

Evidence for a facilitatory role of callosal afferents to the cat motor cortex in the initiation of conditioned bilateral movements.

The effects of selective transection of the rostralmost portion of the corpus callosum, which contains fibres interconnecting the motor cortices of the two hemispheres, on frequency of occurrence and latency of conditioned responses (CRs) in both eyes were examined in seven cats trained to blink in response to a 500-ms tone. A 100-ms air-puff delivered to one eye only (ipsilateral eye) 400 ms after tone onset was used as an unconditioned stimulus. Both before and after callosal lesion, bilateral CRs were the most frequent response pattern. Following callosal lesion, a statistically significant reduction in the percentage of CRs in at least one eye was observed in only two cats. In all seven animals, both before and after callosal lesion, the mean CR latencies of the ipsilateral eye were significantly shorter than those of the contralateral eye. Callosal lesion caused a significant increase in the mean CR latencies of both eyes in all subjects. These results provide evidence that the two hemispheres influence each other in controlling conditioned bilateral blinking by reciprocally exchanging facilitatory signals contributing to initiation of CRs in both eyes.

Training to criterion in eyeblink classical conditioning in Alzheimer's disease, Down's syndrome with Alzheimer's disease, and healthy elderly.

The cholinergic antagonist scopolamine delays acquisition of eyeblink classical conditioning (EBCC) in rabbits and humans, but scopolamine-treated organisms eventually acquire conditioned responses (CRs). Patients with probable Alzheimer's disease (AD) and older adults with Down's syndrome (DS/AD) have disrupted cholinergic systems and perform EBCC very poorly. It was hypothesized that patients with probable AD and DS/AD, like scopolamine-injected organisms, would acquire CRs if given sufficient training. Twelve probable AD patients, 12 DS/AD patients, and 6 healthy elderly control individuals participated in 5 daily 90-trial sessions of EBCC. Fifty-eight percent of the probable AD, 92% of the DS/AD, and 100% of the control participants achieved learning criterion. Probable AD, DS/AD, and control participants had statistically significant increases in the percentage of CRs produced over 5 EBCC sessions. The neural substrate for EBCC was not eliminated in probable AD or DS/AD patients, although the learning mechanism was disrupted.

A nicotinic agonist (GTS-21), eyeblink classical conditioning and nicotinic receptor binding in rabbit brain

The septo-hippocampal cholinergic system is of demonstrated involvement in eyeblink classical conditioning (EBCC). To determine if a nicotinic cholinergic agonist, GTS-21, would facilitate acquisition of EBCC in older rabbits, three doses (0.1, 0.5, 1.0 mg/kg) in sterile saline vehicle and vehicle alone were administered toolder rabbits ( n = 48; mean age= 29.8 months). A control group of vehicle-treated young rabbits ( n = 12; mean age= 3.5 months) was included. Rabbits were conditioned for fifteen 90-trial sessions in the 750 ms delay paradigm with a tone conditioned stimulus and corneal airpuff unconditioned stimulus. Dependent measures of trials to learning criterion, percentage of conditioned responses (CRs) and CR amplitude consistently showed significant improvement in older rabbits treated with 0.5 and 1.0 mg/kg of GTS-21. Acquisition was similar in vehicle-treated young and GTS-treated older rabbits. Vehicle-treated older rabbits conditioned more poorly than vehicle-treated young rabbits. No non-associative learning effects were observed in GTS-21 treated animals. Nicotinic receptor binding was similar in all groups of older rabbits, indicating that GTS-21 administration over a 15-day period did not affect nicotinic receptors. Alzheimer's disease (AD) has been associated with significant loss of nicotinic cholinergic receptors, and patients diagnosed with probable AD are seriously impaired on EBCC. These results demonstrating that the nicotinic agonist, GTS-21, facilitated EBCC in older rabbits suggest that the compound should receive additional investigation for its potential to affect cognition in AD.

The learning capacity of high or low performance rats is related to the hippocampus NMDA receptors

The hippocampal synaptic plasticity of rats with an inborn high (HP) or low (LP) learning capacity to perform in a shuttle ☐ is closely related to their percentage of conditioned responses (Crs). HP rats show less sensitivity to the blocking effect of 2-aminophosphonopentanoic acid (AP5) on the generation of long-term potentiation (LTP) than do LP rats. Results described in the present report are indicative of an increased density of N-methyl- d-aspartate (NMDA) receptors in HP rats compared to control and LP rats. We postulate that the differential pharmacological sensitivity of LTP in these rats is a reflection of this biochemical difference. Also, from these results we suggest that the learning capacity may be related to the density of glutamate NMDA receptors of HP, LP and control rats.

Dihydroergotoxine treatment improves active avoidance performance and increases dopaminestimulated adenylate cyclase activity, in young and aged rats

Dihydroergotoxine (DHET) improved the acquisition of a conditioned avoidance response in both young and old rats, although old animals, reached a lower percentage of conditioned responses than did young rats. DHET dose-dependently reduced locomotor activity in aged animals, while, in young rats, increasing doses caused a triphasic effect, inhibitory at low doses, stimulatory at intermediate doses and again inhibitory at high doses. In young rats, the stimulation of spontaneous movements by DHET was abolished by L-sulpiride; L-sulpiride had no effect in old animals. DHET treatment also potentiated both norepinephrine- and, to a greater extent, dopamine-stimulated frontal cortex adenylate cyclase activity, an effect more pronounced in young than in old rats. These data show that DHET effects both locomotor activity and active avoidance performance in young and aged rats, and modulates dopamine-sensitive adenylate cyclase activity.