Abducens conditioning in in vitro turtle brain stem without cerebellum requires NMDA receptors and involves upregulation of GluR4-containing AMPA receptors

Abstract

Previous work showed that in vitro abducens eyeblink classical conditioning of turtle brain stem-cerebellum preparations involved NMDA-mediated mechanisms and redistribution of GluR4-containing AMPA receptors in the abducens motor nuclei. Since conditioning can be obtained in brain stem preparations without the cerebellum, we examined whether similar mechanisms were involved during conditioning of the brain stem alone. The results showed that conditioning could not be induced in the presence of the NMDA receptor antagonist dl-2-amino-5-phosphonovaleric acid (AP-5) and that abducens nerve conditioned responses, once initiated in normal saline, were significantly attenuated in the presence of AP-5. The effects of AP-5 did not generally depress physiological responsiveness of preparations because some abducens nerve reflexes were not significantly reduced by the compound. GluR4-containing AMPA receptors in the abducens motor nuclei were significantly upregulated and positively correlated with the levels of conditioning similar to that of preparations having an intact cerebellum. Furthermore, increased GluR4 subunits after brain stem conditioning was confirmed by Western blot analysis. These results suggest that NMDA receptor-mediated mechanisms and GluR4 upregulation may mediate in vitro abducens eyeblink classical conditioning and that these mechanisms reside in the brain stem eyeblink circuitry.

Functional Organization of Vestibulo-Ocular Responses in Abducens Motoneurons

“…Here, excitation of all abducens motoneurons is mediated by a mixture of AMPA and NMDA receptors, compatible with the membraneous presence of both receptor molecules (Durand et al, 1987;Durand, 1991;Keifer and Clark, 2003). The absence of a residual NMDA component after blocking the AMPA component reported previously (Straka and Dieringer, 1993) is likely due to the lack of background resting activity from the vestibular sensory periphery in isolated frog brains.…”

Synaptic localization of GluR4-containing AMPARs and Arc during acquisition, extinction, and reacquisition of in vitro classical conditioning

“…Previous studies have demonstrated enhanced synthesis of GluR4 protein and increased localization of GluR4 subunits at synaptic sites corresponding with acquisition and expression of abducens CRs, but not with pseudoconditioning or treatment with the NMDAR antagonist AP-5 which blocks CRs (Keifer, 2001;Keifer & Clark, 2003;Mokin & Keifer, 2004;. In the present study, we have extended these findings to show that the synaptic localization of GluR4-containing AMPARs parallels the expression of abducens CRs during extinction and reacquisition of conditioning.…”

“…In an in vitro model of eyeblink classical conditioning, evidence suggests that synaptic delivery of GluR4-containing AMPARs supports conditioning through NMDAR-mediated mechanisms and MAPK signal transduction (Keifer, 2001;Keifer & Clark, 2003;Mokin & Keifer, 2004;Keifer, Zheng, & Zhu, 2007). In this system, in place of using tone and airpuff stimuli as in behaving animals, weak electrical stimulation of the auditory nerve (the "tone" CS) is paired with strong stimulation of the trigeminal nerve (the "airpuff" US) and results in a neural correlate of conditioned eyeblink responses recorded from the abducens nerve which controls eyeblinks in this species (Keifer, 2003, for a review).…”

Coordinate action of pre- and postsynaptic brain-derived neurotrophic factor is required for AMPAR trafficking and acquisition of in vitro classical conditioning

“…Our initial studies of conditioning used an isolated brain stemcerebellum preparation (Keifer et al, 1995). However, subsequent studies found that an isolated brain stem preparation alone without the cerebellum could acquire robust CRs, although these had a significantly shorter onset latency compared to intact preparations Keifer, 1997, 1999;Keifer and Clark, 2003). These findings are similar to those obtained from rabbits with cerebellar cortex lesions (Perrett et al, 1993).…”