A Fear Memory Engram and Its Plasticity in the Hypothalamic Oxytocin System.

Oxytocin (OT) release by axonal terminals onto the central nucleus of the amygdala exerts anxiolysis. To investigate which subpopulation of OT neurons contributes to this effect, we developed a novel method: virus-delivered genetic activity-induced tagging of cell ensembles (vGATE). With the vGATE method, we identified and permanently tagged a small subpopulation of OT cells, which, by optogenetic stimulation, strongly attenuated contextual fear-induced freezing, and pharmacogenetic silencing of tagged OT neurons impaired context-specific fear extinction, demonstrating that the tagged OT neurons are sufficient and necessary, respectively, to control contextual fear. Intriguingly, OT cell terminals of fear-experienced rats displayed enhanced glutamate release in the amygdala. Furthermore, rats exposed to another round of fear conditioning displayed 5-fold more activated magnocellular OT neurons in a novel environment than a familiar one, possibly for a generalized fear response. Thus, our results provide first evidence that hypothalamic OT neurons represent a fear memory engram.

Peripheral calcium-permeable AMPA receptors regulate chronic inflammatory pain in mice.

α-Amino-3-hydroxy-5-methyl-4-isoxazolepropionic acid-type (AMPA-type) glutamate receptors (AMPARs) play an important role in plasticity at central synapses. Although there is anatomical evidence for AMPAR expression in the peripheral nervous system, the functional role of such receptors in vivo is not clear. To address this issue, we generated mice specifically lacking either of the key AMPAR subunits, GluA1 or GluA2, in peripheral, pain-sensing neurons (nociceptors), while preserving expression of these subunits in the central nervous system. Nociceptor-specific deletion of GluA1 led to disruption of calcium permeability and reduced capsaicin-evoked activation of nociceptors. Deletion of GluA1, but not GluA2, led to reduced mechanical hypersensitivity and sensitization in models of chronic inflammatory pain and arthritis. Further analysis revealed that GluA1-containing AMPARs regulated the responses of nociceptors to painful stimuli in inflamed tissues and controlled the excitatory drive from the periphery into the spinal cord. Consequently, peripherally applied AMPAR antagonists alleviated inflammatory pain by specifically blocking calcium-permeable AMPARs, without affecting physiological pain or eliciting central side effects. These findings indicate an important pathophysiological role for calcium-permeable AMPARs in nociceptors and may have therapeutic implications for the treatment chronic inflammatory pain states.

Activity-dependent potentiation of calcium signals in spinal sensory networks in inflammatory pain states.

The second messenger calcium is a key mediator of activity-dependent neural plasticity. How persistent nociceptive activity alters calcium influx and release in the spinal cord is not well-understood. We performed calcium-imaging on individual cell bodies and the whole area within laminae I and II in spinal cord slices from mice in the naïve state or 24h following unilateral hindpaw plantar injection of complete Freund's adjuvant. Calcium signals evoked by dorsal root stimulation at varying strengths displayed a steep rise and slow decay over 15-20s and increased progressively with both increasing intensity and frequency of stimulation in naïve mice. Experiments with pharmacological inhibitors revealed that both ionotropic glutamate receptors and intracellular calcium stores contributed to maximal calcium signals in laminae I and II evoked by stimulating dorsal roots at 100Hz frequency. Importantly, as compared to naïve mice, we observed that in mice with unilateral hindpaw inflammation, calcium signals were potentiated to 159+/-10% in the ipsilateral dorsal horn and 179+/-8% in the contralateral dorsal horn. In addition to the contribution from NMDA receptors, GluR-A-containing AMPA receptors were found to be critically required for the above changes in spinal calcium signals, as revealed by analysis of genetically modified mouse mutants, whereas intracellular calcium release was not required. Thus, these results suggest that there is an important functional link between calcium signaling in superficial spinal laminae and the development of inflammatory pain. Furthermore, they highlight the importance of GluR-A-containing calcium-permeable AMPA receptors in activity-dependent plasticity in the spinal cord.

Impaired reproductive behavior by lack of GluR-B containing AMPA receptors but not of NMDA receptors in hypothalamic and septal neurons.

The roles of ionotropic glutamate receptors in mammalian reproduction are unknown. We therefore generated mice lacking a major subtype of (S)-alpha-amino-3-hydroxy-5-methyl-isoxazolepropionic acid (AMPA) receptors or all N-methyl-d-aspartate (NMDA) receptors in GnRH neurons and other mainly limbic system neurons, primarily in hypothalamic and septal areas. Male mice without NMDA receptors in these neurons were not impaired in breeding and exhibited similar GnRH secretion as control littermates. However, male mice lacking GluR-B containing AMPA receptors in these neurons were poor breeders and severely impaired in reproductive behaviors such as aggression and mounting. Testis and sperm morphology, testis weight, and serum testosterone levels, as well as GnRH secretion, were unchanged. Contact with female cage bedding failed to elicit male sexual behavior in these mice, unlike in control male littermates. Their female counterparts had unchanged ovarian morphology, had bred successfully, and had normal litter sizes but exhibited pronounced impairments in maternal behaviors such as pup retrieval and maternal aggression. Our results suggest that NMDA receptors and GluR-B containing AMPA receptors are not essential for fertility, but that GluR-B containing AMPA receptors are essential for male and female reproduction-related behaviors, perhaps by mediating responses to pheromones or odorants.