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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

Example histology showing overlap between mPFC terminals anterogradely labeled with td Tomato and retrogradely labeled eyeblink-associated cerebellar neurons. A, Mice that received cerebellar cortex infusions showed retrogradely labeled neurons that extended throughout the anterior–posterior basilar pons (green cells, top 2 images) and into and throughout the RTN (2nd and 3rd images). Substantial overlap between mPFC terminals (magenta) and putative eyeblink-associated cells (green) was observed in the anterior basilar pons (top image, inset 1) and extensively in the RTN (2nd and 3rd images, inset 2). B, Mice that received deep nuclei infusions showed retrogradely labeled neurons (green cells) restricted to the caudal basilar pons (2nd image) and less dense labeling in the more anterior RTN (3rd image). Few if any mPFC terminals were observed in the caudal basilar pons and did not appear to overlap substantially with the labeled pontocerebellar neurons observed there (2nd image, notice the general absence of magenta terminal labeling in inset 3). Overlap was observed the RTN (3rd image, inset 4). Br, Bregma; mcp, middle cerebellar peduncle; ml, medial lemniscus; py, pyramidal tract; scp, superior cerebellar peduncle; MR, median raphe nucleus; RPN, reticular nucleus of the pons.
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Figure 11: Example histology showing overlap between mPFC terminals anterogradely labeled with td Tomato and retrogradely labeled eyeblink-associated cerebellar neurons. A, Mice that received cerebellar cortex infusions showed retrogradely labeled neurons that extended throughout the anterior–posterior basilar pons (green cells, top 2 images) and into and throughout the RTN (2nd and 3rd images). Substantial overlap between mPFC terminals (magenta) and putative eyeblink-associated cells (green) was observed in the anterior basilar pons (top image, inset 1) and extensively in the RTN (2nd and 3rd images, inset 2). B, Mice that received deep nuclei infusions showed retrogradely labeled neurons (green cells) restricted to the caudal basilar pons (2nd image) and less dense labeling in the more anterior RTN (3rd image). Few if any mPFC terminals were observed in the caudal basilar pons and did not appear to overlap substantially with the labeled pontocerebellar neurons observed there (2nd image, notice the general absence of magenta terminal labeling in inset 3). Overlap was observed the RTN (3rd image, inset 4). Br, Bregma; mcp, middle cerebellar peduncle; ml, medial lemniscus; py, pyramidal tract; scp, superior cerebellar peduncle; MR, median raphe nucleus; RPN, reticular nucleus of the pons.

Mentions: Anterogradely labeled tdTomato-expressing mPFC terminal fields were observed in the basilar pons and reticulotegmental nucleus (RTN) in all six injected mice, and showed similar projection patterns (e.g., injection sites: Figs. 10A,B; terminal label: 11A,B; group summary: 13A,B). Densely labeled axons were observed in the medial lemniscus ipsilateral to the mPFC injection site, exiting the white matter bundle and forming a dorsal ring of terminal fields around the medial region of the rostral basilar pons and extending into lateral regions (Figs. 11A, top; summary 13B, left). A small proportion of axons crossed the midline and formed less dense terminal fields in the pons contralateral to the mPFC injection site (Figs. 11A, top; summary 13B, left). Prefrontopontine terminal fields were most dense and stereotyped across the six mice in the anterior third of the basilar pons, with different mice showing unique restricted and less dense terminal fields in the caudal half of the pons (Fig. 13B). However, labeled axons were observed throughout the anterior–posterior extent of the medial lemniscus (Fig. 11A,B). Extensive mPFC terminal labeling was also observed in the RTN, and was most dense near the midline and in the pericentral region, although labeled terminals were observed throughout most of the structure and surrounding regions (Figs. 11A,B; summary 13B). It was unclear, however, whether the source of terminal fields in the RTN included the medial lemniscus, or whether some or all axonal projections may have originated from the adjacent reticular pontine nucleus or from the median raphe, both of which showed extensive mPFC terminal labeling (Fig. 13A,B). The data show that the mPFC of mice projects to the anterior basilar pons and throughout the RTN.


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)

Example histology showing overlap between mPFC terminals anterogradely labeled with td Tomato and retrogradely labeled eyeblink-associated cerebellar neurons. A, Mice that received cerebellar cortex infusions showed retrogradely labeled neurons that extended throughout the anterior–posterior basilar pons (green cells, top 2 images) and into and throughout the RTN (2nd and 3rd images). Substantial overlap between mPFC terminals (magenta) and putative eyeblink-associated cells (green) was observed in the anterior basilar pons (top image, inset 1) and extensively in the RTN (2nd and 3rd images, inset 2). B, Mice that received deep nuclei infusions showed retrogradely labeled neurons (green cells) restricted to the caudal basilar pons (2nd image) and less dense labeling in the more anterior RTN (3rd image). Few if any mPFC terminals were observed in the caudal basilar pons and did not appear to overlap substantially with the labeled pontocerebellar neurons observed there (2nd image, notice the general absence of magenta terminal labeling in inset 3). Overlap was observed the RTN (3rd image, inset 4). Br, Bregma; mcp, middle cerebellar peduncle; ml, medial lemniscus; py, pyramidal tract; scp, superior cerebellar peduncle; MR, median raphe nucleus; RPN, reticular nucleus of the pons.
© Copyright Policy - open-access
Related In: Results  -  Collection

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

Figure 11: Example histology showing overlap between mPFC terminals anterogradely labeled with td Tomato and retrogradely labeled eyeblink-associated cerebellar neurons. A, Mice that received cerebellar cortex infusions showed retrogradely labeled neurons that extended throughout the anterior–posterior basilar pons (green cells, top 2 images) and into and throughout the RTN (2nd and 3rd images). Substantial overlap between mPFC terminals (magenta) and putative eyeblink-associated cells (green) was observed in the anterior basilar pons (top image, inset 1) and extensively in the RTN (2nd and 3rd images, inset 2). B, Mice that received deep nuclei infusions showed retrogradely labeled neurons (green cells) restricted to the caudal basilar pons (2nd image) and less dense labeling in the more anterior RTN (3rd image). Few if any mPFC terminals were observed in the caudal basilar pons and did not appear to overlap substantially with the labeled pontocerebellar neurons observed there (2nd image, notice the general absence of magenta terminal labeling in inset 3). Overlap was observed the RTN (3rd image, inset 4). Br, Bregma; mcp, middle cerebellar peduncle; ml, medial lemniscus; py, pyramidal tract; scp, superior cerebellar peduncle; MR, median raphe nucleus; RPN, reticular nucleus of the pons.
Mentions: Anterogradely labeled tdTomato-expressing mPFC terminal fields were observed in the basilar pons and reticulotegmental nucleus (RTN) in all six injected mice, and showed similar projection patterns (e.g., injection sites: Figs. 10A,B; terminal label: 11A,B; group summary: 13A,B). Densely labeled axons were observed in the medial lemniscus ipsilateral to the mPFC injection site, exiting the white matter bundle and forming a dorsal ring of terminal fields around the medial region of the rostral basilar pons and extending into lateral regions (Figs. 11A, top; summary 13B, left). A small proportion of axons crossed the midline and formed less dense terminal fields in the pons contralateral to the mPFC injection site (Figs. 11A, top; summary 13B, left). Prefrontopontine terminal fields were most dense and stereotyped across the six mice in the anterior third of the basilar pons, with different mice showing unique restricted and less dense terminal fields in the caudal half of the pons (Fig. 13B). However, labeled axons were observed throughout the anterior–posterior extent of the medial lemniscus (Fig. 11A,B). Extensive mPFC terminal labeling was also observed in the RTN, and was most dense near the midline and in the pericentral region, although labeled terminals were observed throughout most of the structure and surrounding regions (Figs. 11A,B; summary 13B). It was unclear, however, whether the source of terminal fields in the RTN included the medial lemniscus, or whether some or all axonal projections may have originated from the adjacent reticular pontine nucleus or from the median raphe, both of which showed extensive mPFC terminal labeling (Fig. 13A,B). The data show that the mPFC of mice projects to the anterior basilar pons and throughout the RTN.

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