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Trace Eyeblink Conditioning in Mice Is Dependent upon the Dorsal Medial Prefrontal Cortex, Cerebellum, and Amygdala: Behavioral Characterization and Functional Circuitry(1,2,3).

Siegel JJ, Taylor W, Gray R, Kalmbach B, Zemelman BV, Desai NS, Johnston D, Chitwood RA - eNeuro (2015)

Bottom Line: To identify the circuitry involved, we made restricted lesions of the medial prefrontal cortex (mPFC) and found that learning was prevented.Anatomical data from these critical regions showed that mPFC and amygdala both project to the rostral basilar pons and overlap with eyelid-associated pontocerebellar neurons.The data further reveal a specific role for the amygdala as providing a conditioned stimulus-associated input to the cerebellum.

View Article: PubMed Central - HTML - PubMed

Affiliation: Center for Learning and Memory, University of Texas at Austin , Austin, Texas 78712.

ABSTRACT
Trace eyeblink conditioning is useful for studying the interaction of multiple brain areas in learning and memory. The goal of the current work was to determine whether trace eyeblink conditioning could be established in a mouse model in the absence of elicited startle responses and the brain circuitry that supports this learning. We show here that mice can acquire trace conditioned responses (tCRs) devoid of startle while head-restrained and permitted to freely run on a wheel. Most mice (75%) could learn with a trace interval of 250 ms. Because tCRs were not contaminated with startle-associated components, we were able to document the development and timing of tCRs in mice, as well as their long-term retention (at 7 and 14 d) and flexible expression (extinction and reacquisition). To identify the circuitry involved, we made restricted lesions of the medial prefrontal cortex (mPFC) and found that learning was prevented. Furthermore, inactivation of the cerebellum with muscimol completely abolished tCRs, demonstrating that learned responses were driven by the cerebellum. Finally, inactivation of the mPFC and amygdala in trained animals nearly abolished tCRs. Anatomical data from these critical regions showed that mPFC and amygdala both project to the rostral basilar pons and overlap with eyelid-associated pontocerebellar neurons. The data provide the first report of trace eyeblink conditioning in mice in which tCRs were driven by the cerebellum and required a localized region of mPFC for acquisition. The data further reveal a specific role for the amygdala as providing a conditioned stimulus-associated input to the cerebellum.

No MeSH data available.


Related in: MedlinePlus

Trace 50–250 CRs were prefrontal-dependent in mice trained to asymptotic performance. A, Unilateral infusions of muscimol (1mm, 100–125 nl) were used to inactivate the mPFC of 5 trained mice (left). Inactivation of the mPFC resulted in reliable decreases in the expression of CRs in trained animals (right graph, red markers; paired t = 4.86, df = 5, p = 0.008), whereas the CR rates observed during control infusions were not different than preinfusion sessions (magenta markers; t = 1.13, p = 0.32). B, Example behavior from muscimol (denoted by red text) and control sessions (magenta text) for two mice. Note the significant decrease in the expression of CRs during muscimol infusion sessions, while behavior during control sessions was less affected by the infusion procedure. Br, Bregma; AGm, medial agranular cortex; AC, anterior cingulate cortex.
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Figure 7: Trace 50–250 CRs were prefrontal-dependent in mice trained to asymptotic performance. A, Unilateral infusions of muscimol (1mm, 100–125 nl) were used to inactivate the mPFC of 5 trained mice (left). Inactivation of the mPFC resulted in reliable decreases in the expression of CRs in trained animals (right graph, red markers; paired t = 4.86, df = 5, p = 0.008), whereas the CR rates observed during control infusions were not different than preinfusion sessions (magenta markers; t = 1.13, p = 0.32). B, Example behavior from muscimol (denoted by red text) and control sessions (magenta text) for two mice. Note the significant decrease in the expression of CRs during muscimol infusion sessions, while behavior during control sessions was less affected by the infusion procedure. Br, Bregma; AGm, medial agranular cortex; AC, anterior cingulate cortex.

Mentions: The lesion experiment indicated that a restricted region of caudal mPFC supports the acquisition of trace eyeblink conditioning in mice. To test whether the same region of mPFC also supports postacquisition expression of trace CRs, we used unilateral infusions of muscimol (100–125 nl) to temporarily inactivate this region of mPFC after training to asymptotic performance (5/8 mice learned, preinfusion mean = 77 ± 4% CR rate). Inactivation resulted in significant decreases in the expression of CRs in 4/5 mice (infusion session mean = 23 ± 11% CR rate, paired t = 4.86, df = 4, p = 0.008w; Fig. A, right; example behavioral sessions given in B, left). As a control to ensure that the infusion procedures alone did not affect the expression of CRs, mice also received infusions of Alexa-conjugated dextran amines dissolved in aCSF (100–125 nl; used to better visualize infusion sites) using the same procedures. Control infusions did not reliably affect the expression of CRs (precontrol infusion mean = 77 ± 6% CR rate, control infusion mean = 61 ± 17% CR rate; paired t = 1.13, df = 4, p = 0.32x; Fig. 7A, right; examples in B, right). Histology revealed that the infusion site of one mouse that showed only a modest decrease in the expression of trace CRs during inactivation was located anterior to the critical region identified in the lesion study (Fig. 7A, left, dashed line indicates anterior border of critical region). The control infusion of a second mouse was unusual in that it resulted in an abolishment of CRs. The histology from this mouse revealed substantial damage at the infusion site, apparently due to a blocked cannula that released a bolus of dextran when positioned. The remaining histology confirmed infusion sites within the critical region of mPFC with minimal damage. The data show that the critical region of mPFC necessary for the acquisition of trace CRs also supports postacquisition expression.


Trace Eyeblink Conditioning in Mice Is Dependent upon the Dorsal Medial Prefrontal Cortex, Cerebellum, and Amygdala: Behavioral Characterization and Functional Circuitry(1,2,3).

Siegel JJ, Taylor W, Gray R, Kalmbach B, Zemelman BV, Desai NS, Johnston D, Chitwood RA - eNeuro (2015)

Trace 50–250 CRs were prefrontal-dependent in mice trained to asymptotic performance. A, Unilateral infusions of muscimol (1mm, 100–125 nl) were used to inactivate the mPFC of 5 trained mice (left). Inactivation of the mPFC resulted in reliable decreases in the expression of CRs in trained animals (right graph, red markers; paired t = 4.86, df = 5, p = 0.008), whereas the CR rates observed during control infusions were not different than preinfusion sessions (magenta markers; t = 1.13, p = 0.32). B, Example behavior from muscimol (denoted by red text) and control sessions (magenta text) for two mice. Note the significant decrease in the expression of CRs during muscimol infusion sessions, while behavior during control sessions was less affected by the infusion procedure. Br, Bregma; AGm, medial agranular cortex; AC, anterior cingulate cortex.
© Copyright Policy - open-access
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC4596016&req=5

Figure 7: Trace 50–250 CRs were prefrontal-dependent in mice trained to asymptotic performance. A, Unilateral infusions of muscimol (1mm, 100–125 nl) were used to inactivate the mPFC of 5 trained mice (left). Inactivation of the mPFC resulted in reliable decreases in the expression of CRs in trained animals (right graph, red markers; paired t = 4.86, df = 5, p = 0.008), whereas the CR rates observed during control infusions were not different than preinfusion sessions (magenta markers; t = 1.13, p = 0.32). B, Example behavior from muscimol (denoted by red text) and control sessions (magenta text) for two mice. Note the significant decrease in the expression of CRs during muscimol infusion sessions, while behavior during control sessions was less affected by the infusion procedure. Br, Bregma; AGm, medial agranular cortex; AC, anterior cingulate cortex.
Mentions: The lesion experiment indicated that a restricted region of caudal mPFC supports the acquisition of trace eyeblink conditioning in mice. To test whether the same region of mPFC also supports postacquisition expression of trace CRs, we used unilateral infusions of muscimol (100–125 nl) to temporarily inactivate this region of mPFC after training to asymptotic performance (5/8 mice learned, preinfusion mean = 77 ± 4% CR rate). Inactivation resulted in significant decreases in the expression of CRs in 4/5 mice (infusion session mean = 23 ± 11% CR rate, paired t = 4.86, df = 4, p = 0.008w; Fig. A, right; example behavioral sessions given in B, left). As a control to ensure that the infusion procedures alone did not affect the expression of CRs, mice also received infusions of Alexa-conjugated dextran amines dissolved in aCSF (100–125 nl; used to better visualize infusion sites) using the same procedures. Control infusions did not reliably affect the expression of CRs (precontrol infusion mean = 77 ± 6% CR rate, control infusion mean = 61 ± 17% CR rate; paired t = 1.13, df = 4, p = 0.32x; Fig. 7A, right; examples in B, right). Histology revealed that the infusion site of one mouse that showed only a modest decrease in the expression of trace CRs during inactivation was located anterior to the critical region identified in the lesion study (Fig. 7A, left, dashed line indicates anterior border of critical region). The control infusion of a second mouse was unusual in that it resulted in an abolishment of CRs. The histology from this mouse revealed substantial damage at the infusion site, apparently due to a blocked cannula that released a bolus of dextran when positioned. The remaining histology confirmed infusion sites within the critical region of mPFC with minimal damage. The data show that the critical region of mPFC necessary for the acquisition of trace CRs also supports postacquisition expression.

Bottom Line: To identify the circuitry involved, we made restricted lesions of the medial prefrontal cortex (mPFC) and found that learning was prevented.Anatomical data from these critical regions showed that mPFC and amygdala both project to the rostral basilar pons and overlap with eyelid-associated pontocerebellar neurons.The data further reveal a specific role for the amygdala as providing a conditioned stimulus-associated input to the cerebellum.

View Article: PubMed Central - HTML - PubMed

Affiliation: Center for Learning and Memory, University of Texas at Austin , Austin, Texas 78712.

ABSTRACT
Trace eyeblink conditioning is useful for studying the interaction of multiple brain areas in learning and memory. The goal of the current work was to determine whether trace eyeblink conditioning could be established in a mouse model in the absence of elicited startle responses and the brain circuitry that supports this learning. We show here that mice can acquire trace conditioned responses (tCRs) devoid of startle while head-restrained and permitted to freely run on a wheel. Most mice (75%) could learn with a trace interval of 250 ms. Because tCRs were not contaminated with startle-associated components, we were able to document the development and timing of tCRs in mice, as well as their long-term retention (at 7 and 14 d) and flexible expression (extinction and reacquisition). To identify the circuitry involved, we made restricted lesions of the medial prefrontal cortex (mPFC) and found that learning was prevented. Furthermore, inactivation of the cerebellum with muscimol completely abolished tCRs, demonstrating that learned responses were driven by the cerebellum. Finally, inactivation of the mPFC and amygdala in trained animals nearly abolished tCRs. Anatomical data from these critical regions showed that mPFC and amygdala both project to the rostral basilar pons and overlap with eyelid-associated pontocerebellar neurons. The data provide the first report of trace eyeblink conditioning in mice in which tCRs were driven by the cerebellum and required a localized region of mPFC for acquisition. The data further reveal a specific role for the amygdala as providing a conditioned stimulus-associated input to the cerebellum.

No MeSH data available.


Related in: MedlinePlus