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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.


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Trace 50–250 CRs were cerebellar-dependent in mice, with no residual noncerebellar behavioral components. A, Infusions of muscimol (1mm, 100–150 nl) were used to inactivate anterior cerebellar regions in 6 trained mice (left). The expression of CRs were completely abolished during infusion sessions in all 6 mice (right graph, red markers; paired t = 28.11, df = 5, p < 0.001), whereas the CR rates observed during control infusions were not different than preinfusion sessions (green markers; t = 1.07, p = 0.34). B, Example behavior from muscimol (denoted by red text) and control sessions (green text) for two mice. Note the complete absence of CRs during muscimol infusion sessions, whereas behavior during control sessions was largely unaffected by the infusion procedure. Br, Bregma; L6, cerebellar cortex lobule 6; L4/5, cerebellar cortex lobule 4/5; DN, dentate nucleus; IN, interpositus nucleus.
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Figure 8: Trace 50–250 CRs were cerebellar-dependent in mice, with no residual noncerebellar behavioral components. A, Infusions of muscimol (1mm, 100–150 nl) were used to inactivate anterior cerebellar regions in 6 trained mice (left). The expression of CRs were completely abolished during infusion sessions in all 6 mice (right graph, red markers; paired t = 28.11, df = 5, p < 0.001), whereas the CR rates observed during control infusions were not different than preinfusion sessions (green markers; t = 1.07, p = 0.34). B, Example behavior from muscimol (denoted by red text) and control sessions (green text) for two mice. Note the complete absence of CRs during muscimol infusion sessions, whereas behavior during control sessions was largely unaffected by the infusion procedure. Br, Bregma; L6, cerebellar cortex lobule 6; L4/5, cerebellar cortex lobule 4/5; DN, dentate nucleus; IN, interpositus nucleus.

Mentions: We tested the cerebellar-dependence of learned responses for trace 50–250 by infusing muscimol into anterior cerebellar regions previously shown to support eyeblink conditioning (at the border of cerebellar lobules 4/5 and 6, and/or the anterior interpositus/dentate nuclei; Fig. 8A, left; McCormick et al., 1982; Yeo et al., 1985a,b; Plakke et al., 2007; Kalmbach et al., 2010a; Gonzalez-Joekes and Schreurs, 2012; Heiney et al., 2014). After mice were trained to asymptotic performance (6/8 implanted mice learned, preinfusion mean = 87 ± 3% CR rate), eyeblink-associated cerebellar regions were inactivated by infusing muscimol (100–150 nl) into the cerebellum 15 min before a training session began. Inactivation resulted in the complete abolishment of CRs in all six cases (infusion session mean = 0 ± 0% CR rate, paired t = 28.11, df = 5, p < 0.001y; Fig. 8A, 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 control infusions of Alexa-conjugated dextran amines dissolved in aCSF (100–150 nl; used to better visualize infusion sites and for anatomical tracing, see below) using the same procedures. The expression of CRs following control infusions was not different from that observed during a typical training session (precontrol infusion mean = 81 ± 6% CR rate, control infusion mean = 77 ± 8% CR rate; paired t = 1.07, df = 5, p = 0.34z; Fig. 8A, right, examples given in B, right). The data show that the entire learned motor response was cerebellar-dependent, and did not include a noncerebellar component as previously observed in mice (Koekkoek et al., 2005; Sakamoto and Endo, 2010).


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 cerebellar-dependent in mice, with no residual noncerebellar behavioral components. A, Infusions of muscimol (1mm, 100–150 nl) were used to inactivate anterior cerebellar regions in 6 trained mice (left). The expression of CRs were completely abolished during infusion sessions in all 6 mice (right graph, red markers; paired t = 28.11, df = 5, p < 0.001), whereas the CR rates observed during control infusions were not different than preinfusion sessions (green markers; t = 1.07, p = 0.34). B, Example behavior from muscimol (denoted by red text) and control sessions (green text) for two mice. Note the complete absence of CRs during muscimol infusion sessions, whereas behavior during control sessions was largely unaffected by the infusion procedure. Br, Bregma; L6, cerebellar cortex lobule 6; L4/5, cerebellar cortex lobule 4/5; DN, dentate nucleus; IN, interpositus nucleus.
© Copyright Policy - open-access
Related In: Results  -  Collection

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

Figure 8: Trace 50–250 CRs were cerebellar-dependent in mice, with no residual noncerebellar behavioral components. A, Infusions of muscimol (1mm, 100–150 nl) were used to inactivate anterior cerebellar regions in 6 trained mice (left). The expression of CRs were completely abolished during infusion sessions in all 6 mice (right graph, red markers; paired t = 28.11, df = 5, p < 0.001), whereas the CR rates observed during control infusions were not different than preinfusion sessions (green markers; t = 1.07, p = 0.34). B, Example behavior from muscimol (denoted by red text) and control sessions (green text) for two mice. Note the complete absence of CRs during muscimol infusion sessions, whereas behavior during control sessions was largely unaffected by the infusion procedure. Br, Bregma; L6, cerebellar cortex lobule 6; L4/5, cerebellar cortex lobule 4/5; DN, dentate nucleus; IN, interpositus nucleus.
Mentions: We tested the cerebellar-dependence of learned responses for trace 50–250 by infusing muscimol into anterior cerebellar regions previously shown to support eyeblink conditioning (at the border of cerebellar lobules 4/5 and 6, and/or the anterior interpositus/dentate nuclei; Fig. 8A, left; McCormick et al., 1982; Yeo et al., 1985a,b; Plakke et al., 2007; Kalmbach et al., 2010a; Gonzalez-Joekes and Schreurs, 2012; Heiney et al., 2014). After mice were trained to asymptotic performance (6/8 implanted mice learned, preinfusion mean = 87 ± 3% CR rate), eyeblink-associated cerebellar regions were inactivated by infusing muscimol (100–150 nl) into the cerebellum 15 min before a training session began. Inactivation resulted in the complete abolishment of CRs in all six cases (infusion session mean = 0 ± 0% CR rate, paired t = 28.11, df = 5, p < 0.001y; Fig. 8A, 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 control infusions of Alexa-conjugated dextran amines dissolved in aCSF (100–150 nl; used to better visualize infusion sites and for anatomical tracing, see below) using the same procedures. The expression of CRs following control infusions was not different from that observed during a typical training session (precontrol infusion mean = 81 ± 6% CR rate, control infusion mean = 77 ± 8% CR rate; paired t = 1.07, df = 5, p = 0.34z; Fig. 8A, right, examples given in B, right). The data show that the entire learned motor response was cerebellar-dependent, and did not include a noncerebellar component as previously observed in mice (Koekkoek et al., 2005; Sakamoto and Endo, 2010).

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