Excitatory Propagation Research Articles

Preceding Administration of Minocycline Suppresses Plastic Changes in Cortical Excitatory Propagation in the Model Rat With Partial Infraorbital Nerve Ligation.

Neuropathic pain is known to be attributable to the injured nerve, a postoperative problem induced by surgery. The infraorbital nerve (ION), a branch of the trigeminal nerve, innervates to the facial and oral regions and conveys somatosensory information to the central nervous system. The partial ligation of ION (pl-ION) is a method to mimic chronic trigeminal neuropathic pain and behavioral abnormality. To counteract induction of such abnormal pain, the effective pharmacological treatment is desired. Although recent studies have revealed the molecular mechanisms regarding chronic pain, estimation of the effectiveness of the pharmacological treatment has not been well-provided especially in the central nervous system so far. Here we examined whether pl-ION induces plastic changes in the cerebral cortex and investigated effects of minocycline on the cortical plastic changes. We performed the pl-ION to Wistar male rats (4–5 weeks old), and confirmed a mechanical nocifensive behavior in response to the mechanical stimulation with von-Frey filaments. The withdrawal threshold to mechanical stimuli of the whisker pad was decreased 1 day (1 d) after pl-ION, which continued up to 14 d after pl-ION, suggesting that pl-ION model rats presented allodynia and enhanced the response sustained at least for 14 d after pl-ION. Next, cerebrocortical activities were evaluated 3 d after pl-ION (3d-pl-ION) by the optical imaging with a voltages-sensitive dye, RH1691, to quantify the response to electrical stimulation of the whisker pad skin, mandibular molar dental pulp, and mentum skin. Electrical stimulation to the whisker pad skin induced smaller excitation in the primary sensory cortex (S1) of 3d-pl-ION in comparison to that in the sham. In contrast, cerebral cortical responses to the mandibular molar dental pulp and mentum skin stimuli increased both in S1, and the secondary somatosensory and insular oral region (S2/IOR) after pl-ION. Administration of minocycline (30 mg/kg/d) from 1 d before to 2 d after pl-ION partially recovered the pl-ION-induced changes in cortical excitation in S1 and S2/IOR in 3d-pl-ION. These results suggest that somatosensory and insular cortical excitation is changed by pl-ION, and the preceding injection of minocycline counteracts the plastic changes in the cortical activities.

Extracellular glucose-dependent IPSC enhancement by leptin in fast-spiking to pyramidal neuron connections via JAK2-PI3K pathway in the rat insular cortex

Leptin is produced in the adipocytes and plays a pivotal role in regulation of energy balance by controlling appetite and metabolism. Leptin receptors are widely distributed in the brain, especially in the hypothalamus, hippocampus, and neocortex. The insular cortex (IC) processes gustatory and visceral information, which functionally correlate to feeding behavior. However, it is still an open issue whether and how leptin modulates IC neural activities. Our paired whole-cell patch-clamp recordings using IC slice preparations demonstrated that unitary inhibitory postsynaptic currents (uIPSCs) but not uEPSCs were potentiated by leptin in the connections between pyramidal (PNs) and fast-spiking neurons (FSNs). The leptin-induced increase in uIPSC amplitude was accompanied by a decrease in paired-pulse ratio. Under application of inhibitors of JAK2-PI3K but not MAPK pathway, leptin did not change uIPSC amplitude. Variance-mean analysis revealed that leptin increased the release probability but not the quantal size and the number of release site. These electrophysiological findings suggest that the leptin-induced uIPSC increase is mediated by activation of JAK2-PI3K pathway in presynaptic FSNs. An in vivo optical imaging revealed that leptin application decreased excitatory propagation in IC induced by electrical stimulation of IC. These leptin-induced effects were not observed under the low energy states: low glucose concentration (2.5 mM) in vitro and one-day-fasting condition in vivo. However, leptin enhanced uIPSCs under application of low glucose with an AMPK inhibitor. These results suggest that leptin suppresses IC excitation by facilitating GABA release in FSN→PN connections, which may not occur under a hunger state.

Trigeminal Nerve Transection-Induced Neuroplastic Changes in the Somatosensory and Insular Cortices in a Rat Ectopic Pain Model.

The primary sensory cortex processes competitive sensory inputs. Ablation of these competitive inputs induces neuroplastic changes in local cortical circuits. However, information concerning cortical plasticity induced by a disturbance of competitive nociceptive inputs is limited. Nociceptive information from the maxillary and mandibular molar pulps converges at the border between the ventral secondary somatosensory cortex (S2) and insular oral region (IOR); therefore, S2/IOR is a suitable target for examining the cortical changes induced by a disturbance of noxious inputs, which often causes neuropathic pain and allodynia. We focused on the plastic changes in S2/IOR excitation in a model of rats subjected to inferior alveolar nerve transection (IANX). Our optical imaging using a voltage-sensitive dye (VSD) revealed that the maxillary molar pulp stimulation-induced excitatory propagation was expanded one to two weeks after IANX at the macroscopic level. At the cellular level, based on Ca2+ imaging using two-photon microscopy, the amplitude of the Ca2+ responses and the number of responding neurons in S2/IOR increased in both excitatory and inhibitory neurons. The in vitro laser scanning photostimulation (LSPS) revealed that Layer II/III pyramidal and GABAergic fast-spiking neurons in S2/IOR received larger excitatory inputs from Layer IV in the IANX models, which supports the findings obtained by the macroscopic and microscopic optical imaging. Furthermore, the inhibitory postsynaptic inputs to the pyramidal neurons were decreased in the IANX models, suggesting suppression of inhibitory synaptic transmission onto excitatory neurons. These results suggest that IANX induces plastic changes in S2/IOR by changing the local excitatory and inhibitory circuits.

Menthol-induced facilitation of cerebrocortical excitatory propagation induced by air puff stimulation of the nasal cavity in the rat: An optical imaging study

Temperature plays a critical role in the sensation of airflow in the nasal mucosa. Neural activities of the ethmoidal nerve, a trigeminal afferent, responding to airflow are suppressed by warm airflow, whereas cold airflow enhances the ethmoidal nerve activities, which is mimicked by application of menthol, a cold-sensitive TRPM8 receptor agonist. However, it has been an open issue how menthol modulates the spatiotemporal profiles of neural activities of somatosensory cortical neurons. In this study, we assessed neural responses to an air puff stimulation (100 ms) to the nasal cavity in the absence or presence of l-menthol using an optical imaging technique with a voltage-sensitive dye in the primary cortex (S1) of urethane-anesthetized rats. A weak air puff application (15 psi) without l-menthol induced neural excitation in a part of the contralateral S1. The air puff stimulation with l-menthol significantly increased the optical signal intensity, expanded the activated area, and shortened the latency, compared to those in the absence of l-menthol. These results suggest that activation of cold-sensitive TRPM8 receptors sharpens airflow sensation in the nasal cavity and expands the receptive field, especially toward the pharynx, which may contribute to enhanced flavor perception.

Experimental tooth movement temporally changes neural excitation and topographical map in rat somatosensory cortex

During orthodontic treatment, binding teeth, may change the topographically organized representation of teeth in the cerebral cortex. To test the hypothesis that experimental tooth movement (ETM) changes the somatotopy of an individual tooth arrangement in the somatosensory cortex, we examined the spatiotemporal features of cortical excitatory propagation in response to mechanical stimulation of the maxillary incisor or molar using optical imaging in late adolescent rats without or with ETM. The ETM models consisted of 1d, 3d, and 7d ETM in which a closed-coil spring was ligated between the maxillary first molar and incisors. In controls, incisor and molar mechanical stimulation evoked excitation in the rostral and dorsocaudal regions of the primary somatosensory cortex (S1), respectively. In addition, the secondary somatosensory cortex and insular oral region (S2/IOR) were also activated. Incisor stimulation-induced excitatory regions in S1 of 3d and 7d ETM shifted without changing the maximum excitatory area or peak amplitude; the incisor stimulation-responding region moved toward the dorsocaudal region, which responded to molar stimulation in the control. This shift in excitatory region was not observed in 1d ETM. One day after removal of the coil spring that was attached for 6 days, the excitatory region shift in S1 was recovered to the control region. On the other hand, 1d ETM exhibited facilitation of the excitatory area and peak amplitude upon molar stimulation, and the facilitation of excitatory propagation disappeared in 3d and 7d ETM. These results may explain the clinical finding that abnormal sensation temporally occurs during orthodontic treatment.

Somatotopic Organization and Temporal Characteristics of Cerebrocortical Excitation in Response to Nasal Mucosa Stimulation With and Without an Odor in the Rat: An Optical Imaging Study

Nasal mucosa has roles in warming and humidifying inspired air and is highly sensitive to mechanical stimuli. Moreover, the upper part of the nasal mucosa expresses olfactory receptors processing olfactory information. Although the somatosensory map of the face in the primary (S1) and secondary (S2) somatosensory cortices is clearly documented, the map of the nasal mucosa and the effect of odors on their activities are largely unknown. This study aimed to identify the cortical regions in S1 and their temporal features in response to somatosensory stimulation of the nasal mucosa using an optical imaging technique in urethane-anesthetized rats. An air puff application response to nasal mucosa first occurred in a part of contralateral S1 and subsequently, spread toward the rostrally and ventrally adjacent sites. Upper pharynx stimulation initially activated this rostrally expanded site and the excitatory propagation from the initially activated region toward ventral region likely represented S2. Signal intensity and activated area increased dependent on air pressure. Nasal tip stimulation initially excited S1 region caudally adjacent to that of nasal mucosa. Moreover, the amplitude of S1 excitation was similar between air puff stimulation with and without an odor, amyl acetate. In contrast to contralateral S1, air puff stimulation with the odor showed a faint optical signal increase in the ipsilateral piriform cortex. These results suggest that somatosensory information from the nasal mucosa and skin, and upper pharynx are processed in spatially continuous regions of S1, and interaction between somatosensory and olfactory systems is relatively small in contralateral S1.

Ablation of C-fibers decreases quantal size of GABAergic synaptic transmission in the insular cortex

The primary sensory cortex exhibits neuroplastic changes responding to sensory disturbances, and GABAergic synaptic transmission plays a critical role in the regulation of plasticity. The insular cortex (IC) integrates orofacial nociceptive signals conveyed via myelinated Aδ- and unmyelinated C-fibers. However, it has been unknown whether a disturbance of nociceptive inputs, such as a deletion of the peripheral nerves, alters GABAergic local circuit in IC. The present study elucidated GABAergic synaptic transmission in the model rat whose C-fibers were ablated by capsaicin injection 1–2 days after birth. In vivo optical imaging revealed that capsaicin-treated rats showed a facilitative excitatory propagation in IC responding to dental pulp stimulation. Whole-cell patch-clamp recording from pyramidal neurons (Pyr) demonstrated that capsaicin-treated rats showed the smaller amplitude of miniature inhibitory postsynaptic currents (IPSCs) than sham-treated rats without changing the frequency. Furthermore, replacement of extracellular Ca2+ to Sr2+, which induces an asynchronous release of neurotransmitters in the quantal size, induced a smaller amplitude of asynchronous unitary IPSCs recorded from fast-spiking GABAergic interneuron to Pyr connections in capsaicin-treated rats than sham-treated rats. These results suggest that capsaicin treatment depresses IPSCs via a postsynaptic mechanism. To confirm this possibility, the variance-mean analysis of unitary IPSCs was employed and we found that quantal size of GABAergic synaptic transmission was smaller in capsaicin-treated rats than in sham-treated rats. These results suggest that ablation of C-fibers induces plastic changes in GABAergic synaptic transmission by decreasing postsynaptic GABAA receptor-mediated conductance, which is a possible mechanism of the facilitative excitation in IC of capsaicin-treated rats.

Opioid subtype- and cell-type-dependent regulation of inhibitory synaptic transmission in the rat insular cortex

The insular cortex (IC) plays a principal role in the regulation of pain processing. Although opioidergic agonists depress cortical excitatory synaptic transmission, little is known about opioidergic roles in inhibitory synaptic transmission. In the IC, the opioid receptors differentially regulate the excitatory propagation: agonists of the mu (MOR), delta (DOR), and kappa (KOR) exhibit suppressive, facilitative, and little effects, respectively. Thus, we aimed to examine the effects of opioid receptor agonists on unitary inhibitory postsynaptic currents (uIPSCs) in the IC. Pyramidal and GABAergic neurons in the rat IC were recorded by a multiple whole-cell patch-clamp technique. [D-Ala2,N-Me-Phe4,Gly5-ol]-Enkephalin acetate salt (DAMGO), an MOR agonist, reduced uIPSC amplitude by 74% in fast-spiking GABAergic interneuron (FS)→FS connections without a significant effect on FS→pyramidal cell (Pyr) connections. These effects of DAMGO were also observed in non-FS→FS and non-FS→Pyr connections: DAMGO reduced the uIPSC amplitude in non-FS→FS but not in non-FS→Pyr connections. DAMGO-induced depression of uIPSCs was blocked by the MOR antagonist, D-Phe-Cys-Tyr-D-Trp-Arg-Thr-Pen-Thr-NH2. The DOR agonist, [D-Pen2,5]-Enkephalin hydrate (DPDPE), reduced uIPSC amplitude by 39% in FS→FS and by 49% in FS→Pyr connections, which was antagonized by the DOR antagonist, naltrindole. However, DPDPE had little effect on non-FS→FS/Pyr connections. (±)-trans-U-50488 methanesulfonate salt (U50488), a KOR agonist, had little effect on uIPSC in FS→FS/Pyr connections. These results suggest that MOR-induced uIPSC depression in FS→FS and non-FS→FS, but not FS→Pyr and non-FS→Pyr connections, results in the depression of excitatory propagation in the IC, which may be an underlying mechanism of the powerful analgesic effects of MOR agonists.

Opposite effects of mu and delta opioid receptor agonists on excitatory propagation induced in rat somatosensory and insular cortices by dental pulp stimulation

The insular cortex (IC) contributes to nociceptive information processing. IC neurons express opioid receptors, including the mu (MOR), kappa (KOR), and delta (DOR) subtypes. Opioidergic agonists suppress excitatory synaptic transmission in the cerebral cortex. In addition, morphine injection into the IC reduces responses to noxious thermal stimuli. However, the mechanisms of the opioid-dependent modulation of cortical excitation at the macroscopic level, which bridge the cellular and behavioral findings, have remained unknown. The present in vivo optical imaging study aimed to examine the effects of the agonists of each subtype on cortical excitatory propagation in the IC and the neighboring cortices, the primary (S1) and secondary somatosensory (S2) areas. To assess the opioidergic effects on the cortical circuits, we applied electrical stimulation to the maxillary 1st molar pulp, which induced excitation in the ventral part of S1 and the S2/insular oral region (IOR). The initial excitatory response was observed 10–14ms after stimulation, and then excitation propagated concentrically. DAMGO (10–100μM), an MOR agonist, suppressed the amplitude of cortical excitation and shrank the maximum excitation areas in S1 and S2/IOR. In contrast, 10–100μM DPDPE, a DOR agonist, increased the amplitude of excitation and expanded the area of maximum excitation. U50488 (10–100μM), a KOR agonist, had little effect on cortical excitation. These results suggest that MOR-induced suppression of excitatory propagation in the IC is an underlying mechanism of the powerful analgesic effects of MOR agonists. In contrast, DOR may play a minor role in suppressing acute pain.

Sequential Changes in Cortical Excitation during Orthodontic Treatment

Cortical excitation responding to periodontal ligament (PDL) stimulation is observed in the rat primary somatosensory (S1), secondary somatosensory, and insular oral region of the cortex (S2/IOR), which are considered to process somatosensation, including nociception. Our previous studies have demonstrated that excitatory propagation induced by PDL stimulation is facilitated in S1 and S2/IOR 1 d after experimental tooth movement (ETM), and tetanic stimulation of IOR induces long-term potentiation of cortical excitatory propagation consistently. These findings raise the possibility that ETM induces neuroplastic changes, and as a result, facilitation of cortical excitation would be sustained for weeks. However, no information is available about the temporal profiles of the facilitated cortical responses. We estimated PDL stimulation-induced cortical excitatory propagation in S1 and S2/IOR of rats by optical imaging 1 to 7 d after ETM of the maxillary first molar. ETM models showed facilitated cortical excitatory propagation in comparison with controls and sham groups 1 d after ETM, but the facilitation gradually recovered to the control level 3 to 7 d after ETM. Sham groups that received wire fixation without orthodontic force tended to enhance cortical responses, although the differences between controls and sham groups were almost insignificant. We also examined the relationship between cortical responses and expression of inflammatory cytokines, interleukin (IL)–1β and tumor necrosis factor (TNF)–α, in PDL of the first molar. The peak amplitude of optical signals responding to PDL stimulation tended to be increased in parallel to the number of IL-1β and TNF-α immunopositive cells, suggesting that, at least in part, the enhancement of cortical responses is induced by PDL inflammation. These findings suggest that ETM-induced facilitation of cortical excitatory propagation responding to PDL stimulation 1 d after ETM recovers to the control level within a week. The time course of the facilitated cortical responses is comparable to that of pain and discomfort induced by clinical orthodontic treatments.

Anatomical and electrophysiological mechanisms for asymmetrical excitatory propagation in the rat insular cortex: In vivo optical imaging and whole-cell patch-clamp studies

The insular cortex (IC) integrates limbic information from the amygdala and hypothalamic nucleus to multimodal sensory inputs, including visceral, gustatory, and somatosensory information. However, the functional framework of excitation in the IC is still unknown. We performed optical imaging and single pyramidal neuronal staining using a whole-cell patch-clamp technique in urethane-anesthetized rats to elucidate the precise anatomical and physiological features of IC pyramidal neurons, which regulate cortical information processing via their horizontal connections. Optical imaging revealed that electrical stimulation of the granular (GI) or dysgranular (DI) IC elicited characteristic excitatory propagations along the rostrocaudal axis parallel to the rhinal fissure, with a preference toward the rostral direction. Spatial patterns of the dendrites and axons of layer II/III pyramidal cells in the DI/GI support these functional features of excitation; for example, rostrocaudal axonal arbors tend to extend with a rostral directional preference. The mean length of the axons from the soma to the farthest site rostrally was ∼50% longer than that of the caudal length. Pyramidal cells in the DI/GI exhibited spontaneous membrane oscillation in the UP and DOWN states. Similarly to the evoked signals obtained by optical imaging, repetitive electrical stimulation of the caudal IC ∼1 mm away from the recorded cells (five pulses at 50 Hz) induced the summation of evoked excitatory postsynaptic potentials during the DOWN state and profound inhibitory postsynaptic potentials during the UP state. Clarification of the excitation feature with its cellular basis provides new clues about the functional mechanisms of the asymmetric propagation of neural activities in the IC.

GABAB receptors accentuate neural excitation contrast in rat insular cortex

Synaptic transmission mediated by metabotropic GABA receptors, GABAB receptors, regulates physiological functions of cerebrocortical local circuits. It is, however, still unknown how GABAB receptors regulate excitatory propagation at more macroscopic level. We performed in vivo optical imaging to investigate the spatio-temporal profiles of GABAB receptor-mediated regulation of excitatory propagation in anesthetized rat insular cortex (IC). Repetitive electrical stimulation (a sequence of 10 pulses at 50 Hz) of the dysgranular IC (DI), a part of gustatory cortex (GC), elicited excitatory propagation along the rhinal fissure. Tonic activation of GABAB receptors by application of baclofen suppressed the optical signal amplitude to the early pulses in the sequence (first to third stimuli), typically in the rostral GC (rGC). In contrast, optical signal amplitude to later pulses was enhanced by baclofen in both the rGC and caudal GC (cGC). Baclofen reduced the area of excitation during the early pulses in the sequence but not during later pulses. Application of CGP 52432, which blocked GABAB receptor-mediated tonic and phasic inhibition, slightly suppressed optical responses to early pulses (though not to the first pulse), whereas it enhanced responses to later pulses, especially in the dorsolateral orbital cortex (DLO). Decay amplitude of the response to the first pulse was reversed to a large rise in amplitude by the GABAA receptor antagonist bicuculline. The decay amplitude was enhanced by CGP 52432 and reversed to a small rise by baclofen. This suggests that GABAB receptor activation reduced postsynaptic GABAA receptor activation indirectly via inhibition of presynaptic GABA release. Optical responses induced by DLO stimulation were reduced by pre-stimulation of the cGC 180 ms before DLO stimulation, which was blocked by CGP 52432. These results suggest that tonic and phasic activation of GABAB receptors cooperatively enhances the contrast of neural excitation at a level of millimeters.

Spatiotemporal dynamics of long-term potentiation in rat insular cortex revealed by optical imaging

Long-term potentiation (LTP) of the gustatory cortex (GC), a part of the insular cortex (IC) around the middle cerebral artery, is a key process of gustatory learning and memory, including conditioned taste aversion learning. The rostral (rGC) and caudal GC (cGC) process different tastes; the rGC responds to hedonic and the cGC responds to aversive tastes. However, plastic changes of spatial interaction of excitatory propagation between the rGC and cGC remain unknown. The present study aimed to elucidate spatiotemporal profiles of excitatory propagation, induced by electrical stimulation (five train pulses) of the rGC/cGC before and after LTP induction, using in vivo optical imaging with a voltage-sensitive dye. We demonstrated that tetanic stimulation of the cGC induced long-lasting expansion of the excitation responding to five train stimulation of the cGC, and an increase in amplitude of optical signals in the IC. Excitatory propagation after LTP induction spread preferentially toward the rostral IC: the length constant ( λ) of excitation, obtained by fitting optical signals with a monoexponential curve, was increased to 121.9% in the rostral direction, whereas λ for the caudal, dorsal, and ventral directions were 48.9%, 44.2%, and 62.5%, respectively. LTP induction was prevented by pre-application of D-APV, an NMDA receptor antagonist, or atropine, a muscarinic receptor antagonist, to the cortical surface. In contrast, rGC stimulation induced only slight LTP without direction preference. Considering the different roles of the rGC and cGC in gustatory processing, these characteristic patterns of LTP in the GC may be involved in a mechanism underlying conversion of palatability.

Pilocarpine-induced status epilepticus causes acute interneuron loss and hyper-excitatory propagation in rat insular cortex

Recent clinical studies have shown that the insular cortex (IC) is involved in temporal lobe epilepsy and suggested that the IC mediates spreading of epileptic activity from the temporal lobe, including the hippocampus and amygdala, to the frontal cortex. However, little is known about anatomical and physiological features of the IC in models of temporal lobe epilepsy. The present study evaluated the distribution pattern of GABAergic interneurons, especially parvalbumin (PV)- and somatostatin (SS)-immunopositive neurons, and excitatory propagation pattern in the IC of rats 4–7 days and 2 months after pilocarpine-induced status epilepticus (4–7 d and 2 m post-SE rats, respectively). The number of PV-immunopositive neuron profiles in the agranular IC (AI) significantly decreased by 24.6% and 41.5% in 7 d and 2 m post-SE rats, respectively. The dysgranular and granular IC (DI+GI) exhibited only 5.2% loss of PV-immunopositive neurons in 7 d post-SE rats, while 2 m post-SE rats showed 30.4% loss of PV-immunopositive neurons. There was no significant change of the SS-immunopositive neuron profile numbers in the AI and DI+GI of 7 d and 2 m post-SE rats. The regions with decreased numbers of PV-immunopositive neuron profiles overlapped with those where many degenerating cells were detected by Fluoro-Jade B staining. The area of excitatory propagation responding to electrical stimulation of the caudal AI was expanded in 4–7 d post-SE rats, and excitation frequently propagated to the frontal cortex including the motor cortex. Optical signals in the AI of 4–7 d post-SE rats were larger in amplitude than those of controls. In contrast to the AI, the DI of 4–7 d post-SE rats showed similar excitatory propagation pattern and amplitude to that of controls. These results suggest that the region-specific loss of PV-immunopositive neurons occurred in the AI 4–7 d after pilocarpine-induced status epilepticus, which may play an important role in facilitating excitatory propagation in the IC.

Optical recordings reveal the differential distribution of glutamatergic transmission in the cochlear and vestibular nuclei in the newborn mouse brainstem

ObjectiveUsing optical imaging in brainstem slices, we studied evoked responses in the mouse cochlear (CN) and vestibular (VN) nuclei. MethodsThe use of optical imaging allowed us to visualize the spatiotemporal patterns of excitatory propagation in the CN and VN. Optical recordings can differentiate excitatory propagation in the ventral CN(VCN) from that in the dorsal CN(DCN). Furthermore, we assessed the relative distribution of NMDA and non–NMDA receptors in these regions using the glutamate antagonists APV (NMDA receptor antagonist) and CNQX (non–NMDA receptor antagonist) in mouse brainstem slices during postnatal days 1 to 3. ResultsThe average sensitivity to APV was 99.1% in the VCN, 76.0% in the DCN and 64.9% in the VN. The average sensitivity to CNQX was 0.9% in the VCN, 24.0% in the DCN and 35.1% in the VN. These results indicate that the effect of APV dominated in the VCN, while both APV and CNQX were effective antagonists in the DCN and VN. In the VN, the distribution of APV– and CNQX–sensitive cells was almost completely uniform. However, in the DCN the distribution of APV– and CNQX–sensitive cells was highly complex. The area that was more sensitive to CNQX was located in the superficial layer of the DCN while the area with a higher sensitivity to APV was located progressively in the deep layer. ConclusionThis optical recording data suggests that there is a differential distribution of NMDA and non–NMDA receptor mediated neurotransmission in the VCN, DCN and VN.

Spatiotemporal dynamics of excitation in rat insular cortex: intrinsic corticocortical circuit regulates caudal-rostro excitatory propagation from the insular to frontal cortex

The insular cortex (IC), composing unique anatomical connections, receives multi-modal sensory inputs including visceral, gustatory and somatosensory information from sensory thalamic nuclei. Axonal projections from the limbic structures, which have a profound influence on induction of epileptic activity, also converge onto the IC. However, functional connectivity underlying the physiological and pathological roles characteristic to the IC still remains unclear. The present study sought to elucidate the spatiotemporal dynamics of excitatory propagation and their cellular mechanisms in the IC using optical recording in urethane-anesthetized rats. Repetitive electrical stimulations of the IC at 50 Hz demonstrated characteristic patterns of excitatory propagation depending on the stimulation sites. Stimulation of the granular zone of the IC (GI) and other surrounding cortices such as the motor/primary sensory/secondary sensory cortices evoked round-shaped excitatory propagations, which often extended over the borders of adjacent areas, whereas excitation of the agranular and dysgranular zones in the IC (AI and DI, respectively) spread along the rostrocaudal axis parallel to the rhinal fissure. Stimulation of AI/DI often evoked excitation in the dorsolateral orbital cortex, which exhibited spatially discontinuous topography of excitatory propagation in the IC. Pharmacological manipulations using 6,7-dinitroquinoxaline-2,3(1H,4H)-dione (DNQX), a non-NMDA receptor antagonist, D-2-amino-5-phosphonovaleric acid (D-APV), an NMDA receptor antagonist, and bicuculline methiodide, a GABAA receptor antagonist, indicate that excitatory propagation was primarily regulated by non-NMDA and GABAA receptors. Microinjection of lidocaine or incision of the supragranular layers of the rostrocaudally middle part of excitatory regions suppressed excitation in the remote regions from the stimulation site, suggesting that the excitatory propagation in the IC is largely mediated by cortical local circuits. These features of excitatory propagation in the AI/DI, that is the propagation along the rostrocaudal axis with less propagation in the ventro-dorsal direction, may play an important role for transmitting neural excitation arising from the limbic structures to the frontal and orbital cortices.

Differential regulation of synaptic transmission by adrenergic agonists via protein kinase A and protein kinase C in layer V pyramidal neurons of rat cerebral cortex

Activation of α 1- and β-adrenoceptors modulates excitatory neural transmission in the cerebral cortex in opposite manners. Our in vitro optical imaging study using a voltage sensitive dye has revealed that an α 1-adrenoceptor agonist, phenylephrine, suppresses the excitatory propagation evoked by stimulation of the white matter, whereas a β-adrenoceptor agonist, isoproterenol, tends to potentiate the excitatory propagation especially in the deeper layers. The present study aimed to explore what kind of second messengers are involved in noradrenergic modulation of synaptic transmission by using intracellular recording in rat cerebrocortical slice preparation. Evoked excitatory postsynaptic potentials (eEPSPs) were recorded from regular spiking and bursting pyramidal neurons in layer V, which generate single and complex action potentials in response to a short (5 ms) depolarizing current pulse injection, respectively. Application of phenylephrine attenuated eEPSPs, and prazosin, an α 1-adrenoceptor antagonist, precluded the phenylephrine-induced suppression of eEPSPs. The EPSPs suppression by phenylephrine was blocked by pre-application of a protein kinase C (PKC) inhibitor, chelerythrine, whereas a PKC activator, phorbol 12-myristate 13-acetate (phorbol ester), mimicked the effect of phenylephrine. On the other hand, application of isoproterenol enhanced eEPSPs, and propranolol, a β-adrenoceptor antagonist, precluded the excitatory effect of isoproterenol on eEPSPs. The membrane permeant analog of cyclic-3′,5′-AMP (cAMP), N 6,2′-O-dibutyryl-AMP (db-cAMP), mimicked the facilitatory effect of isoproterenol. Isoproterenol-induced enhancement of eEPSPs was promoted by pre-application of 4-(3-butoxy-4-methoxybenzyl)-2-imidazolidinone (Ro 20–1724), a cAMP-specific phosphodiesterase inhibitor. A selective protein kinase A inhibitor, N-[2-( p-bromocinnamylamino)ethyl]-5-soquinolinesulfonamide (H-89), inhibited the excitatory effect by isoproterenol. There was no significant difference in the effects of adrenergic agonists/antagonists and protein kinase activators/inhibitors between regular spiking and bursting neurons in layer V. Thus, it is likely that the suppressive effect on eEPSPs by activation of α 1-adrenoceptors was mediated by protein kinase C, and excitatory effect by activation of β-adrenoceptors was mediated by cAMP/protein kinase A cascade in layer V pyramidal neurons.

Optical mapping of neural responses and their gamma-aminobutyric acid-ergic inhibitory effects in the auditory brainstem of early postnatal mice.

gamma-Aminobutyric acid (GABA)ergic neurons play important tropic and modulatory roles in the auditory pathway, especially in the early stage between postnatal Days 0 and 5. The effects of GABA and GABAa receptor antagonist were observed in this experimental study. Numerous histological and electrophysiological studies have been performed on the contribution of GABA to the auditory pathway; however, the spatio-temporal patterns of excitatory propagation and the relationships between GABA receptor and excitatory propagation have yet to be reported. Using an optical recording technique and a voltage-sensitive dye, the spatio-temporal patterns of excitatory propagation were observed in the auditory brainstem slices of early postnatal mice. A bath containing 50 microM GABA was applied, which largely inhibited the excitatory activities along the vestibulocochlear pathway. Bicuculline methiodide (BMI), a competitive antagonist against GABAa receptor, partially reversed the effects of GABA on the optical signals. Bath application of BMI alone helped to facilitate the depolarization course and its effect was apparent as an enlargement of the depolarized region from the cochlear nucleus and vestibular nucleus to some adjacent brainstem nuclei, as well as enhancing the amplitude of changes in the optical signals. The experimental results seem to suggest that GABAa receptors are widely distributed in an early postnatal auditory brainstem. GABA exhibited a greater modulating effect in the adjacent brainstem nuclei, which are involved in complex information processes, than that observed in the modulating primary auditory pathway. In the present experiment, significant GABAergic contributions to the optical recordings in the auditory brainstem were observed.

Multiple-site optical recording of mouse brainstem evoked by vestibulocochlear nerve stimulation

We used optical imaging to investigate the mouse cochlear and vestibular nucleus in brainstem slices using a voltage-sensitive dye, RH 155. As a result, the spatiotemporal patterns of excitatory propagation were shown. These optical signals consisted of two components consisting of a spike-like fast signal and a long-lasting slow signal. All responses were abolished by tetrodotoxin. The slow signals were eliminated under a Ca 2+-free solution. In addition, synaptic fatigue was also observed. The present study indicated the feasibility of optical recording for visually revealing the synaptic transmission in both the vestibular and cochlear nucleus.

Selective suppression of horizontal propagation in rat visual cortex by norepinephrine.

The release of norepinephrine in the cerebral cortex from axon terminals of locus coeruleus neurons was suggested to be involved in the control of attention. Accumulating data indicate that the responses of cortical neurons are varied when norepinephrine is applied iontophoretically in the vicinity of the cells being recorded. However, it is not known how the pattern of excitatory propagation is modified when norepinephrine is applied over a wide area in the visual cortex. By applying optical imaging to rat visuocortical slices, we found a new mode of norepinephrine action; a prominent suppression of the horizontal propagation in layers II/III. This action of norepinephrine was confirmed by the simultaneous recording of field potentials from multiple sites by use of a multi-electrode dish. Furthermore, our electrophysiological recordings showed that this norepinephrine action is exerted through suppression of excitatory neural transmission and enhancement of inhibitory transmission to the pyramidal neurons in these layers. Because the release of norepinephrine in the visual cortex is regulated by the level of attention, the neural basis of visual attention may relate partially to the suppression of the integration of visual information by norepinephrine resulting in a state-dependent restructuring of the receptive field.