Korean J Physiol Pharmacol.  2017 Sep;21(5):487-493. 10.4196/kjpp.2017.21.5.487.

Inhibition of anterior cingulate cortex excitatory neuronal activity induces conditioned place preference in a mouse model of chronic inflammatory pain

• Kang SJ
• Kim S
• Lee J
• Kwak C
• Lee K
• Zhuo M
• Kaang BK

• KMID: 2388694
• DOI: http://doi.org/10.4196/kjpp.2017.21.5.487

Abstract

The anterior cingulate cortex (ACC) is known for its role in perception of nociceptive signals and the associated emotional responses. Recent optogenetic studies, involving modulation of neuronal activity in the ACC, show that the ACC can modulate mechanical hyperalgesia. In the present study, we used optogenetic techniques to selectively modulate excitatory pyramidal neurons and inhibitory interneurons in the ACC in a model of chronic inflammatory pain to assess their motivational effect in the conditioned place preference (CPP) test. Selective inhibition of pyramidal neurons induced preference during the CPP test, while activation of parvalbumin (PV)-specific neurons did not. Moreover, chemogenetic inhibition of the excitatory pyramidal neurons alleviated mechanical hyperalgesia, consistent with our previous result. Our results provide evidence for the analgesic effect of inhibition of ACC excitatory pyramidal neurons and a prospective treatment for chronic pain.

Keyword

Anterior cingulate cortex; Conditioned place preference; Optogenetics; Pain

MeSH Terms

Animals
Chronic Pain
Gyrus Cinguli*
Hyperalgesia
Interneurons
Mice*
Neurons*
Optogenetics
Prospective Studies
Pyramidal Cells

Figure

• 

  Fig. 1 Experimental schematic of the conditioned place preference testThe CFA-injected mice were first freely exposed to the apparatus for 15 min (Day 1, Pre-test). Next, the mice were conditioned with light in the compartment with less preference, and without light in the compartment with more preference (Days 2~7, conditioning). After 6 days of conditioning, the mice were assessed for changed preference for the conditioning compartments (Day 8, Post-test).

• 

  Fig. 2 Conditioned place preference in the CaMKII-eNpHR group.(A) Sample image of eNpHR expression in the ACC. (B) Result of exploration time in the less preferred compartment on Day 1 (Pre-test). There was no significant difference between the EYFP (31.62±1.74%, n=10) and eNpHR (28.39±0.98%; n=14) groups in exploration time (p=0.0978, unpaired t-test). (C) Result of the time spent in the conditioned compartment on Day 8 (Post-test, normalized to the pre-test exploration time). There was a significant difference between the EYFP (0.97±0.06, n=10) and eNpHR (1.25±0.10; n=14) groups (p=0.0441, unpaired t-test).

• 

  Fig. 3 Conditioned place preference in the PV-ChR2 group.(A) Sample image of ChR2 expression in the ACC. (B) Result of exploration time in the less preferred compartment on Day 1 (Pre-test). There was no significant difference between the cre(−) (29.50±3.23%, n=4) and cre(+) (27.11±1.90%, n=8) groups in exploration time (p=0.5110, unpaired t-test). (C) Result of the time spent in the conditioned compartment on Day 8 (Post-test, normalized to the pre-test exploration time). There was no significant difference between the cre(−) (1.06±0.03, n=4) and cre(+) (1.16±0.12, n=8) groups (p=0.7481, unpaired t-test).

• 

  Fig. 4 Results of immunohistochemistry and electronic von Frey test in the CaMKII-hM4Di group.(A) Experimental schematic of the electronic von Frey test. Mice were allowed to recover from brain surgery for about 3 weeks. On Day 0, CFA was injected into the hind paw of each mouse, after determining the baseline response to the electronic von Frey apparatus. On Day 3, CNO was administered to each mouse 40 min before the experiment, and the response to the von Frey apparatus was measured. (B) Mechanical threshold responses to the electronic von Frey test on Days 0 and 3. There was no significant difference between the mCherry (4.81±0.09 g; n=20) and hM4Di (4.94±0.10 g; n=19) groups in the baseline responses (p=0.507, post hoc Bonferroni test) on Day 0. After CNO administration to both groups, the mechanical threshold showed a significant difference between groups (hM4Di [4.70±0.16; n=19], mCherry [3.76±0.19; n=20], p<0.001, post hoc Bonferroni test) on Day 3. The mCherry group displayed significantly different mechanical thresholds between days (Day 0 vs. Day 3, p<0.001). The mechanical threshold of the hM4Di group was not significantly different between days (Day 0 vs. Day 3, p=0.15). Repeated-measures two-way ANOVA with post hoc Bonferroni correction demonstrated a significant effect of CNO on Day 3 (p<0.001), significant effect of DREADD (p=0.002), significant difference between days (p <0.001), and a significant effect of interaction between two factors (Virus x Time, p=0.0013). (C) Images of the ACC of the mCherry (left) and hM4Di (right) groups after immunohistochemistry. (D) The ratio of activated neurons labeled by c-Fos among mCherry(+) neurons in the mCherry group was significantly different from that in the hM4Di group (p=0.0002, unpaired t-test).

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