Elsevier

Neuroscience

Volume 163, Issue 1, 29 September 2009, Pages 190-201
Neuroscience

Cellular Neuroscience
Research Paper
Cannabinoid signaling in inhibitory autaptic hippocampal neurons

https://doi.org/10.1016/j.neuroscience.2009.06.004Get rights and content

Abstract

Depolarization-induced suppression of excitation and inhibition (DSE/DSI) appears to be an important form of short-term retrograde neuronal plasticity involving endocannabinoids (eCBs), the activation of presynaptic cannabinoid CB1 receptors, and the suppression of neurotransmitter release. Using murine autaptic hippocampal cultures, we have distinguished five populations of autaptic inhibitory neurons that exhibit differential cannabinoid responses, including three temporally distinct forms of DSI. One remaining population responded to cannabinoids but did not have DSI while a fifth had neither DSI nor cannabinoid responses. Of the two chief candidate eCBs, 2-AG reversibly inhibited inhibitory post synaptic currents (IPSCs) while anandamide did so irreversibly, the latter's action inconsistent with a role as a bona fide eCB mediator of DSI. The duration of depolarization necessary to elicit the two most prominent forms of DSI (effective dose (ED-50) ∼210, ∼280 ms) was far less than for autaptic DSE. However the nearly identical concentration response for 2-AG to inhibit excitatory postsynaptic currents (EPSCs) and IPSCs indicates that this difference is not due to differential cannabinoid receptor sensitivity. Interestingly, of the two populations exhibiting prominent DSI, one had a substantially faster recovery time course both after DSI and 2-AG, this despite being cultured under identical conditions. Several enzymes have been proposed to play a role in 2-AG breakdown, presumably determining the time course of DSI: fatty acid amide hydrolase (FAAH), cyclooxygenase-2 (COX-2), monoacyl glycerol lipase (MGL), and α/β-hydrolase domains 6 and 12 (ABHD6 and ABHD12). We tested the impact on DSI duration by blockers of FAAH, COX-2, MGL and ABHD6. Notably, the population with slow DSI was regulated only by MGL, whereas the fast DSI population was regulated by both MGL and COX-2. This suggests that the faster DSI time course may occur as a result of the concerted action of multiple enzymes, which may represent a more general mechanism for regulation of the duration of different forms of DSI and DSE.

Section snippets

Culture preparation

All procedures used in this study were approved by the Animal Care Committees of Indiana University and the University of Washington and conform to the guidelines of the National Institutes of Health on the Care and Use of Animals. Experiments were designed in such a way as to minimize the number of animals used and their suffering. Mouse hippocampal neurons isolated from the CA1–CA3 region were cultured on microislands as described previously (Furshpan et al 1976, Bekkers and Stevens 1991).

Five classes of inhibitory autaptic hippocampal neurons based on their response to stimulation and cannabinoids

IPSCs are easily distinguishable from excitatory postsynaptic currents (EPSCs) by virtue of their visibly slower decay time course (e.g. Fig. 1B, decay time constant for representative EPSC (t½): 4.0 ms; for IPSC: 17.1 ms). Because they represent approximately 10% of the total autaptic population, and are difficult to identify reliably by eye, these neurons were initially avoided/discarded. However in the process of recording from several of these neurons it quickly became apparent that they

Discussion

We have identified four classes of cannabinoid-responsive inhibitory autaptic hippocampal neurons, three of them exhibiting temporally distinct forms of autDSI. As with autDSE, a single autaptic neuron is therefore capable of functionally expressing both the pre- and post-synaptic components of retrograde eCB signaling. This preparation offers an architecturally simple model for detailed investigation of the mechanisms of distinct variants of DSI and how they contrast to those for DSE.

Our

Conclusion

Because they represent less than 10% of cultured hippocampal neurons, and so are encountered too infrequently to yield datasets of sufficient size, inhibitory autaptic hippocampal neurons are generally avoided. The neurons can nonetheless be classified into five populations based on their responses to three stimuli: cannabinoids, a DSI-evoking three second depolarization and a high frequency stimulus. The distinction of such populations among neurons grown under otherwise identical conditions

Acknowledgments

The work was supported by IU LMIC, by NIH grant DA11322 and the Lilly Endowment, Inc.

References (34)

  • T.P. Dinh et al.

    Brain monoglyceride lipase participating in endocannabinoid inactivation

    Proc Natl Acad Sci U S A

    (2002)
  • D. Fegley et al.

    Characterization of the fatty-acid amide hydrolase inhibitor URB597: effects on anandamide and oleoylethanolamide deactivation

    J Pharmacol Exp Ther

    (2004)
  • T.F. Freund et al.

    Interneurons of the hippocampus

    Hippocampus

    (1996)
  • E.J. Furshpan et al.

    Chemical transmission between rat sympathetic neurons and cardiac myocytes developing in microcultures: evidence for cholinergic, adrenergic, and dual-function neurons

    Proc Natl Acad Sci U S A

    (1976)
  • L.L. Glickfeld et al.

    Distinct timing in the activity of cannabinoid-sensitive and cannabinoid-insensitive basket cells

    Nat Neurosci

    (2006)
  • M. Isokawa et al.

    Retrograde endocannabinoid regulation of GABAergic inhibition in the rat dentate gyrus granule cell

    J Physiol

    (2005)
  • I. Katona et al.

    Presynaptically located CB1 cannabinoid receptors regulate GABA release from axon terminals of specific hippocampal interneurons

    J Neurosci

    (1999)
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