Yoji Osako

Oxytocin enhances presynaptic and postsynaptic glutamatergic transmission between rat olfactory bulb neurones in culture.

Although oxytocin (OT) within the olfactory bulb has been implicated in maternal behaviour and olfactory recognition, the cellular mechanisms of action remain to be clarified. We examined the effects of OT on glutamatergic spontaneous excitatory postsynaptic currents (sEPSCs) in cultured granule cells with the use of whole-cell patch-clamp recordings. OT reversibly increased both the frequency and amplitude of sEPSCs. The effects of OT on sEPSCs were blocked by the selective OT receptor antagonist desGly-NH(2)(9),d (CH(2))(5)-[Thy(Me)(2),Thr(4)]-ornithine vasotocin. OT had no detectable effect, however, on high voltage-activated Ca2+ currents in mitral/tufted cells, suggesting that OT acts presynaptically on step(s) in the release process downstream from calcium influx. OT augmented the membrane current in granule cells evoked by exogenous application of glutamate, indicating a postsynaptic site of action. These results indicate that OT facilitates sEPSCs in granule cells by both pre- and postsynaptic mechanisms.

Juvenile social experience regulates central neuropeptides relevant to emotional and social behaviors

Stressful social experiences during early-life can increase the risk of developing neuropsychiatric disorders associated with anxiety, mood, and personality. Early neglect also alters peripheral arginine vasopression (AVP) and oxytocin (OXT). We hypothesized that a lack of social stimuli should adversely affect developmental AVP and OXT systems. To test this idea, we examined changes of central AVP- and OXT-immunoreactive (ir) cell number as well as its related behaviors in socially isolated rats. Animals were weaned at 23 days of age, divided into group- or isolation-reared conditions, and maintained for at least 2 weeks. At 38-48 days of age, animals were sacrificed for immunohistochemistry, or used for two behavioral tests: elevated plus-maze test and social recognition test. The results from immunohistochemistry showed that isolation-reared males have decreased AVP-ir cells in the paraventricular nucleus hypothalamus (PVH), medial parvicellular part, ventral zone, and that isolation-reared females have decreased OXT-ir cells in the PVH, medial parvicellular part, dorsal zone, when compared with group-reared counterparts. The results from behavioral assessment showed that isolation-reared animals have difficulty with social recognition, and that isolation-reared males, but not females, have anxiogenic profile. The present study demonstrates that post-weaning social isolation results in decrease of male AVP-ir cells and female OXT-ir cells in the PVH parvocellular divisions, and supports the idea that juvenile social environment may play a critical role in neuronal and behavioral development.

A mental retardation gene, motopsin/neurotrypsin/prss12, modulates hippocampal function and social interaction

Motopsin is a mosaic serine protease secreted from neuronal cells in various brain regions, including the hippocampus. The loss of motopsin function causes nonsyndromic mental retardation in humans and impairs long‐term memory formation in Drosophila. To understand motopsin’s function in the mammalian brain, motopsin knockout (KO) mice were generated. Motopsin KO mice did not have significant deficits in memory formation, as tested using the Morris water maze, passive avoidance and Y‐maze tests. A social recognition test showed that the motopsin KO mice had the ability to recognize two stimulator mice, suggesting normal social memory. In a social novelty test, motopsin KO mice spent a longer time investigating a familiar mouse than wild‐type (WT) mice did. In a resident–intruder test, motopsin KO mice showed prolonged social interaction as compared with WT mice. Consistent with the behavioral deficit, spine density was significantly decreased on apical dendrites, but not on basal dendrites, of hippocampal pyramidal neurons of motopsin KO mice. In contrast, pyramidal neurons at the cingulate cortex showed normal spine density. Spatial learning and social interaction induced the phosphorylation of cAMP‐responsive element‐binding protein (CREB) in hippocampal neurons of WT mice, whereas the phosphorylation of CREB was markedly decreased in mutant mouse brains. Our results indicate that an extracellular protease, motopsin, preferentially affects social behaviors, and modulates the functions of hippocampal neurons.

Modulation of dendrodendritic interactions and mitral cell excitability in the mouse accessory olfactory bulb by vaginocervical stimulation

When female mice are mated, they form a memory to the pheromonal signal of their male partner. The neural changes underlying this memory occur in the accessory olfactory bulb, depend upon vaginocervical stimulation at mating and involve changes at the reciprocal synapses between mitral and granule cells. However, the action of vaginocervical stimulation on the reciprocal interactions between mitral and granule cells remains to be elucidated. We have examined the effects of vaginocervical stimulation on paired‐pulse depression of amygdala‐evoked field potentials recorded in the external plexiform layer of the accessory olfactory bulb (AOB) and the single‐unit activity of mitral cells antidromically stimulated from the amygdala in urethane‐anaesthetized female mice. Artificial vaginocervical stimulation reduced paired‐pulse depression (considered to be due to feedback inhibition of the mitral cell dendrites from the granule cells via reciprocal dendrodendritic synapses) recorded in the AOB external plexiform layer. As would be expected from this result, vaginocervical stimulation also enhanced the spontaneous activity of a proportion of the mitral cells tested. These results suggest that vaginocervical stimulation reduces dendrodendritic feedback inhibition to mitral cells and enhances their activity.

Changes in calcitonin gene-related peptide expression following joint immobilization in rats

Long-term immobilization by casting can occasionally cause pathologic pain states in the immobilized side. The underlying neurophysiological mechanisms of immobilization-related pain are not well understood. For this reason, we specifically examined changes of calcitonin gene-related peptide (CGRP) expression in the dorsal root ganglion (DRG), spinal dorsal horn and posterior nuclei (cuneate nuclei) in a long-term immobilization model following casting for 5 weeks. A plastic cast was wrapped around the right limb from the forearm to the forepaw to keep wrist joint at 90 degrees of flexion. In this model, CGRP in immobilized (ipsilateral) side was distributed in larger DRG neurons compared with contralateral side, even though the number of CGRP-immunoreactive (CGRP-IR) neurons did not differ. Spinal laminae III-V, not laminae I-II in ipsilateral side showed significantly high CGRP expression relative to contralateral side. CGRP expression in cuneate nuclei was not significantly different between ipsilateral and contralateral sides. Long-term immobilization by casting may induce phenotypic changes in CGRP expression both in DRG and spinal deep layers, and these changes are partly responsible for pathological pain states in immobilized side.

Enzymatic properties and localization of motopsin (PRSS12), a protease whose absence causes mental retardation

Motopsin (PRSS12) is a mosaic protease expressed in the central nervous system. Truncation of the human motopsin gene causes nonsyndromic mental retardation. Understanding the enzymatic properties and localization of motopsin protein in the central nervous system will help identify the molecular mechanism by which the loss of motopsin function causes mental retardation. Recombinant motopsin showed amidolytic activity against the synthetic substrate benzyloxycarbonyl-l-phenylalanyl-l-arginine 4-methyl-coumaryl-7-amide. Motopsin activated the single-chain tissue plasminogen activator precursor and exhibited gelatinolytic activity. This enzymatic activity was inhibited by typical serine protease inhibitors such as aprotinin, leupeptin, and (4-amidinophenyl) methanesulfonyl fluoride. Immunocytochemistry using anti-motopsin IgG revealed that both human and mouse motopsin proteins were distributed in discrete puncta along the dendrites and soma as well as axons in cultured hippocampal neurons. In the limbic system, including the cingulate and hippocampal pyramidal neurons and piriform cortex, high level of motopsin protein was expressed at postnatal day 10, but a very low level at 10-week-old mice. Motopsin and tissue plasminogen activator were co-expressed in the cingulate pyramidal neurons at postnatal day 10 and were distributed along dendrites of cultured pyramidal neurons. In cranial nuclei, a moderate level of motopsin protein was detected independently on the developmental stage. Our results suggest that motopsin has multiple functions, such as axon outgrowth, arranging perineuronal environment, and maintaining neuronal plasticity, partly in coordination with other proteases including tissue plasminogen activator.

Oxytocin depresses spontaneous γ-aminobutyric acid-ergic inhibitory postsynaptic currents in cultured mitral cells of the rat olfactory bulb by a presynaptic mechanism

Oxytocin (OT) modulation of synaptic transmission between olfactory bulb neurones has been implicated in the induction of maternal behaviour, but the mechanism of action is unknown. We examined the action of OT on gamma-aminobutyric acid(A) (GABA(A)) receptor-mediated spontaneous inhibitory postsynaptic currents (sIPSCs) in cultured mitral/tufted (M/T) cells with the use of whole-cell patch-clamp recordings. OT reversibly reduced the frequency of sIPSCs without affecting the amplitudes. The effect of OT on sIPSCs was mimicked by the OT receptor agonist [Thr(4), Gly(7)]-OT in a reversible manner and blocked by the OT receptor antagonist desGly-NH(2)(9), d(CH(2))(5)-[Tyr(Me)(2), Thr(4)]-ornithine-vasotocin. OT has no effect, however, on the membrane currents evoked by exogenous application of GABA. These results demonstrate that OT depresses GABA(A) receptor-mediated sIPSCs in M/T cells by a presynaptic mechanism.

A mental retardation gene, motopsin/prss12, modulates cell morphology by interaction with seizure-related gene 6.

A serine protease, motopsin (prss12), plays a significant role in cognitive function and the development of the brain, since the loss of motopsin function causes severe mental retardation in humans and enhances social behavior in mice. Motopsin is activity-dependently secreted from neuronal cells, is captured around the synaptic cleft, and cleaves a proteoglycan, agrin. The multi-domain structure of motopsin, consisting of a signal peptide, a proline-rich domain, a kringle domain, three scavenger receptor cysteine-rich domains, and a protease domain at the C-terminal, suggests the interaction with other molecules through these domains. To identify a protein interacting with motopsin, we performed yeast two-hybrid screening and found that seizure-related gene 6 (sez-6), a transmembrane protein on the plasma membrane of neuronal cells, bound to the proline-rich/kringle domain of motopsin. Pull-down and immunoprecipitation analyses indicated the interaction between these proteins. Immunocytochemical and immunohistochemical analyses suggested the co-localization of motopsin and sez-6 at neuronal cells in the developmental mouse brain and at motor neurons in the anterior horn of human spinal cords. Transient expression of motopsin in neuro2a cells increased the number and length of neurites as well as the level of neurite branching. Interestingly, co-expression of sez-6 with motopsin restored the effect of motopsin at the basal level, while sez-6 expression alone showed no effects on cell morphology. Our results suggest that the interaction of motopsin and sez-6 modulates the neuronal cell morphology.

Partner Loss in Monogamous Rodents: Modulation of Pain and Emotional Behavior in Male Prairie Voles

Objective Pain is modulated by psychosocial factors, and social stress–induced hyperalgesia is a common clinical symptom in pain disorders. To provide a new animal model for studying social modulation of pain, we examined pain behaviors in monogamous prairie voles experiencing partner loss. Methods After cohabitation with novel females, males (n = 79) were divided into two groups on the basis of preference test scores. Half of the males of each group were separated from their partner (loss group), whereas the other half remained paired (paired group). Thus, males from both groups experienced social isolation. Open field tests, plantar tests, and formalin tests were then conducted on males to assess anxiety and pain-related behaviors. Results Loss males showing partner preferences (n = 20) displayed a significant increase in anxiety-related behavior in the open-field test (central area/total distance: 13.65% [1.58%] for paired versus 6.45% [0.87%] for loss; p < .001), a low threshold of thermal stimulus in the plantar test (withdrawal latencies: 9.69 [0.98] seconds for paired versus 6.15 [0.75] seconds for loss; p = .037), and exacerbated pain behaviors in the formalin test (total number of lifts: 40.33 [4.46] for paired versus 54.42 [1.91] for loss; p = .042) as compared with paired males (n = 20). Thermal thresholds in the plantar test significantly correlated with anxiety-related behavior in the open-field test (r = 0.64). No such differences were observed in the males that did not display partner preferences (r = 0.15). Conclusions Results indicate that social bonds and their disruption, but not social housing without bonding followed by isolation, modulate pain and emotion in male prairie voles. The prairie vole is a useful model for exploring the neural mechanisms by which social relationships contribute to pain and nociceptive processing in humans.

A single prolonged stress paradigm produces enduring impairments in social bonding in monogamous prairie voles.

Traumatic events such as natural disasters, violent crimes, tragic accidents, and war, can have devastating impacts on social relationships, including marital partnerships. We developed a single prolonged stress (SPS) paradigm, which consisted of restraint, forced swimming, and ether anesthesia, to establish an animal model relevant to post-traumatic stress disorder. We applied a SPS paradigm to a monogamous rodent, the prairie vole (Microtus ochrogaster) in order to determine whether a traumatic event affects the establishment of pair bonds. We did not detect effects of the SPS treatment on anhedonic or anxiety-like behavior. Sham-treated male voles huddled with their partner females, following a 6day cohabitation, for a longer duration than with a novel female, indicative of a pair bond. In contrast, SPS-treated voles indiscriminately huddled with the novel and partner females. Interestingly, the impairment of pair bonding was rescued by oral administration of paroxetine, a selective serotonin reuptake inhibitor (SSRI), after the SPS treatment. Immunohistochemical analyses revealed that oxytocin immunoreactivity (IR) was significantly decreased in the supraoptic nucleus (SON), but not in the paraventricular nucleus (PVN), 7days after SPS treatment, and recovered 14days after SPS treatment. After the presentation of a partner female, oxytocin neurons labeled with Fos IR was significantly increased in SPS-treated voles compared with sham-treated voles regardless of paroxetine administration. Our results suggest that traumatic events disturb the formation of pair bond possibly through an interaction with the serotonergic system, and that SSRIs are candidates for the treatment of social problems caused by traumatic events. Further, a vole SPS model may be useful for understanding mechanisms underlying the impairment of social bonding by traumatic events.