Canna~Fangled Abstracts

Novel Song-Stimulated Dendritic Spine Formation and Arc/Arg 3.1 Expression in Zebra Finch Auditory Telencephalon are Disrupted by Cannabinoid Agonism.

By October 19, 2013No Comments
 [Epub ahead of print]

pm2Novel Song-Stimulated Dendritic Spine Formation and Arc/Arg 3.1 Expression in Zebra Finch Auditory Telencephalon are Disrupted by Cannabinoid Agonism.

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Department of Pharmacology and Toxicology, Brody School of Medicine, East Carolina University, Greenville, North Carolina 27834.

Abstract

Cannabinoids are well-established to alter processes of sensory perception; however neurophysiological mechanisms responsible remain unclear. Arc, an immediate-early gene (IEG) product involved in dendritic spine dynamics and necessary for plasticity changes such as long-term potentiation, is rapidly induced within zebra finch caudal medial nidopallium (NCM) following novel song exposure, a response that habituates after repeated stimuli. Arc appears unique in its rapid postsynaptic dendritic expression following excitatory input. Previously, we found that vocal development-altering cannabinoid treatments are associated with elevated dendritic spine densities in motor- (HVC) and learning-related (Area X) song regions of zebra finch telencephalon. Given Arc’s dendritic morphological role, we hypothesized that cannabinoid-altered spine densities may involve Arc-related signaling. To test this, we examined the ability of the cannabinoid agonist WIN55212-2 (WIN) to: (1) acutely disrupt song-induced Arc expression; (2) interfere with habituation to auditory stimuli and; (3) alter dendritic spine densities in auditory regions. We found that WIN (3mg/kg) acutely reduced Arc expression within both NCM and Field L2 in an antagonist-reversible manner. WIN did not alter Arc expression in thalamic auditory relay Nucleus Ovoidalis (Ov), suggestingcannabinoid signaling selectively alters responses to auditory stimulation. Novel song stimulation rapidly increased dendritic spine densities within auditory telencephalon, an effect blocked by WIN pretreatments. Taken together, cannabinoid inhibition of both Arc induction and its habituation to repeated stimuli, combined with prevention of rapid increases in dendritic spine densities, implicates cannabinoid signaling in modulation of physiological processes important to auditory responsiveness and memory.
© 2013 Published by Elsevier B.V.

KEYWORDS:

Arc, Arc/Arg 3.1, Auditory, Auditory Field L2, Cannabinoid, Caudal Medial Nidopallium, Dendritic spines, L2, Learning, MAP2, Memory, NCM, Nucleus Ovoidalis, Ov, SR, SR141716A, Sensory, Songbird, WIN, WIN55212-2, activity-regulated cytoskeleton-associated protein (also known as Arg3.1), microtubule-associated protein 2

PMID:

 24134952
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elsevierAbbreviations

  • Arc, activity-regulated cytoskeleton-associated protein (also known as Arg3.1);
  • WIN, WIN55212-2;
  • SR,SR141716A;
  • L2, Auditory Field L2;
  • NCM, Caudal Medial Nidopallium;
  • Ov, Nucleus ovoidalis;
  • MAP2,microtubule-associated protein 2

Keywords

  • Dendritic spine;
  • Cannabinoid;
  • Auditory;
  • Sensory;
  • Songbird;
  • Arc/Arg3.1;
  • Learning;
  • Memory

Figures and tables from this article:

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Fig. 1. Schematic representation of the avian auditory brain regions studied. The thalamic region nucleus ovoidalis (Ov) relays primary auditory sensory information to thalamorecipient Field L2 (L2, shown in light gray), which projects to caudal medial neostriatum (NCM, shown in darker gray).
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Fig. 2. (A) Representative Western blot of the soluble fraction of protein isolated from male NCM brain homogenates. Similar to other species, the anti-Arc antibody labels a single predominant band of about 55 kDa (left lane). Staining is eliminated by preabsorption with 20 μM of the immunizing peptide (right lane). (B) Novel song stimulation increases Arc protein levels in zebra finch caudal telencephalon that contains auditory regions NCM and Field L2 (compare Silence to Song). This stimulation was prevented by pretreatment with the cannabinoid agonist WIN (WIN+Song). Blots were re-probed with an antibody directed against β-actin as an internal control. (C) Summary of the relative intensities of Arc:β-actin bands. Shown are means ±SEM of five pooled experiments. Novel song stimulation (Song) increased Arc expression over that of silence-exposed controls (Silence). WIN treatment prior to novel song exposure (Song+WIN) reduced Arc expression. Asterisk indicates significant difference from the Silence group, dagger indicates difference from Song group (p<0.05, 1-tailed t-test).
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Fig. 3. Double immunofluorescence labeling of Arc protein with dendritically-associated MAP2 (630×,1000×), visualized via confocal laser scanning microscopy. In an effort to capture Arc protein expression at its maximum level, brains of animals exposed to novel song were used for these experiments. Images demonstrating Arc distribution alone (green), and colocalized with MAP2 (yellow, image overlays) show both cellular and dendritic distribution of protein. Bar=20 µm.
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Fig. 4. Immunohistochemical staining of NCM (solid boxes in (A), (D), (G)) and Field L2 (dashed boxes in (A), (D), (G)) with Arc antibody. Animals were exposed to either vehicle (D)–(F) or WIN pretreatment (G)–(I) 30 min before novel song exposure. Unstimulated control animals (A–C) received only silence. At 1000× magnification, novel song stimulation increases cellular and neuropil Arc expression within both NCM (E) and Field L2 (F). (J)–(K) Optical densities of NCM (J) and Field L2 (K) Arc expression after novel song exposure, and after WIN pretreatment. Novel song stimulation increases Arc expression by about 90%, but only by about 20% after WIN exposure. Bar in (G) 300 µm. Bar in (H)–(I) 30 µm.
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Fig. 5. Optical densities (G) and immunohistochemical staining (A)–(F) of Arc expression within Ov in unstimulated controls, after acute novel song exposure, and after pretreatment with WIN. The absence of cannabinoid-mediated effects on Arc expression within this thalamic regions in contrast with those observed in telencephalon, supporting brain region selectivity of responsiveness. Scale bar in (E) 300 µm. Scale bar in (F) 30 µm.
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Fig. 6. Birds were exposed to the same 30 min song for 3 days. On days 1 and 2, birds received WIN (3 mg/kg), the CB1-selective antagonist SR (6 mg/kg) and WIN, or vehicle injections 30 min before song stimulation. On day 3, all groups received vehicle injections prior to song exposure, were euthanized 90 min afterward and brains processed for anti-Arc immunolabeling (A)–(O). (G)–(I) Habituation to novel song-induced Arc expression occurred in the vehicle treated group as evidenced by a return to baseline levels. WIN exposure disrupts habituation (J)–(L) however, and this disruption is reversed with the antagonist SR (M)–(O).
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Fig. 7. (A) Representative high power image of an NCM spiny neuron used for analysis. Bar=30 μm. (B)–(D) Representative Golgi–Cox impregnated dendrites included in analysis of dendritic spine densities within (B) unstimulated control, (C) 30 min of novel song exposure and (D) cannabinoid-pretreated groups (3 mg/kg WIN 30 min prior to song exposure). (E) Novel song stimulation results in significantly increased dendritic spine densities within NCM. Note that this effect was produced rapidly after acute auditory exposure (animals were killed 90 min following song exposure). Cannabinoid pretreatment (30 min prior to song exposure) with 3 mg/kg WIN prevents rapid increases in spine densities.
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Copyright © 2013 Published by Elsevier B.V.

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