The antidiabetic drug metformin has been found to have beneficial effects in various neurological disorders; however, the molecular mechanisms underlying these effects remain unclear. Here we report that metformin protects neuronal cells from quinolinic acid (QUIN)-induced excitotoxicity. For this, we pretreated N18D3 neuronal cells with metformin prior to QUIN for 24 h. We found that pretreating the cells with metformin significantly improved cell survival rate in a concentration-dependent manner and reduced apoptotic cell death, as revealed by a MTT assay and DAPI staining, respectively. Calcium imaging using fluo-4 showed that metformin (100 µM) inhibited the intracellular calcium increase that was induced by QUIN. In addition, mRNA expression of pro-apoptotic genes, p21 and Bax, was decreased and of anti-apoptotic genes, Bcl-2 and Bcl-xl, was increased with metformin treatment compared to QUIN-induced cells. The immunoreactivity of phosphorylated ERK1/2 was elevated in cells treated with metformin, indicating the ERK1/2 signaling pathway in the neuroprotective effects of metformin in QUIN-induced cell death. Collectively, our data demonstrates that metformin exerts its neuroprotective effects by inhibiting intracellular calcium increases, allowing it to regulate ERK1/2 signaling and modulate cell survival and death genes.
Apoptosis ; Calcium ; Cell Death ; Cell Survival ; Genes, bcl-2 ; Metformin ; Nervous System Diseases ; Neurons ; Neuroprotection ; Neuroprotective Agents ; Quinolinic Acid ; RNA, Messenger

In the present study, the vestibularly evoked activity of inferior olive (IO) neurons was examined to investigate the vertical vestibular information transmitted through the vestibulo-olivo-cerebellar climbing fiber pathway. The extracellular recording was made in 74 neurons of the IO of cats, while animals were sinusoidally rotated. Most of vestibularly activated IO neurons responded to the vertical rotation (roll) test and were found in or near the beta subnuclei (IObeta). The vestibular IO neurons were activated, when the animal was rotated to the side contralateral to the recording site. In contrast to the observation that the gain of responses of yaw sensitive cells (YSC) was not changed by the rotation frequency, that of the roll-sensitive cells (RSC) decreased as the rotation frequency was increased. Regardless of RSC or HSC, IO neurons showed the tendency of phase-lag in their responses. The alternating excitatory and inhibitory phases of responses of RSC were dependent on the direction of head orientation, the characterstics of which are the null response plane (NRP) and the optimal response plane (ORP). The analysis based on the NRP of RSC showed that vestibular inputs from the ipsilateral anterior semicircular canal induced the NRP of the RSC response at about 45 degree counterclockwise to the longitudinal axis of the animal, and that those inputs were distributed to RSC in the rostral part of IObeta. On the other hand, those from the posterior semicircular canal were related with the NRP at about 45 degree clockwise and with the caudal part of the IObeta. These results suggest that IO neurons receive and encode the vestibular information, the priority of which seems to be the vertical component of the body movement rather than the horizontal ones.
Animals ; Axis, Cervical Vertebra ; Cats ; Hand ; Head ; Neurons* ; Olea* ; Semicircular Canals*

The present study was primarily carried out to characterize the properties of the spinomesencephalic tract (SMT) neurons that project from the upper cervical spinal segments to the midbrain. It was also investigated whether these neurons received convergent afferent inputs from other sources in addition to cervical inputs. Extracellular single unit recordings were made from neurons antidromically activated by stimulation of midbrain. Recording sites were located in lamina IapprxVIII of C1apprxC3 segments of spinal cord. Receptive field (RF) and response properties to mechanical stimulation were studied in 71 SMT neurons. Response profiles were classified into six groups: complex (Comp, n = 9), wide dynamic range (WDR, n = 16), low threshold (LT, n = 5), high threshold (HT, n = 6), deep/tap (Deep, n = 10), and nonresponsive (NR, n = 25). Distributions of stimulation and recording sites were not significantly different between SMT groups classified upon their locations and/or response profiles. Mean conduction velocity of SMT neurons was 16.7 +/- 1.28 m/sec. Conduction velocities of SMTs recorded in superficial dorsal horn (SDH, n = 15) were significantly slower than those of SMTs recorded in deep dorsal horn (DDH, n = 18), lateral reticulated area (LRA, n = 21), and intermediate zone and ventral horn (IZ/HV n = 15). Somatic RFs for SMTs in LRA and IZ/VH were significantly larger than those in SDH and DDH. Five SMT units (4 Comps and 1 HT) had inhibitory somatic RFs. About half (25/46) of SMT units have their RFs over trigeminal dermatome. Excitabilities of 5/12 cells and 9/13 cells were modulated by stimulation of ipsilateral phrenic nerve and vagus nerve, respectively. These results suggest that upper cervical SMT neurons are heterogenous in their function by showing a wide range of variety in location within the spinal gray matter, in response profile, and in convergent afferent input.
Animals ; Cats* ; Horns ; Mesencephalon* ; Neurons* ; Phrenic Nerve ; Spinal Cord ; Vagus Nerve

In spite of abundant anatomical evidences for the fiber connection between vestibular nuclei and inferior olivary (IO) complex, the transmission of vestibular information through the vestibulo- olivo-cerebellar climbing fiber pathway has not been physiologically established. The aims of the present study were to investigate whether there are IO neurons specifically responding to horizontal rotation and also in which subregions of IO complex these vestibularly-activated neurons are located. The extracellular recording was made in 68 IO neurons and responses of 46 vestibularly-activated cells were analyzed. Most of the vestibularly-activated IO neurons responded to signals of vertical rotation (roll), while a small number (13/46) of recorded cells were activated by horizontal canal signal (yaw). Regardless of yaw-sensitive or roll-sensitive, vestibular IO neurons were excited, when the animal was rotated to the side contralateral to the recording side. The gain and excitation phase were very similar to otolithic or vertical-canal responses. Histologic identification of recording sites showed that most of vestibular IO neurons were located in beta subnucleus. Electrical stimulation of a HSC evoked an inhibitory effect on the excitability of the ipsilateral IO neurons. These results suggest that IO neurons mainly in the beta subnucleus receive vestibular signals from semicircular canals and otolithic organs, encode them, and transmit vestibular information to the cerebellum.
Animals ; Cerebellum ; Electric Stimulation ; Neurons* ; Olea* ; Otolithic Membrane ; Semicircular Canals* ; Vestibular Nuclei

We examined the neurotoxic effects of 3 glutathione (GSH) depletors, buthionine sulfoximine (BSO), diethyl maleate (DEM) and phorone, under the presence of trolox, cycloheximide (CHX), pyrrolidine dithiocarbamate (PDTC) or MK-801 in primary mouse cortical cell cultures. All three depletors induced neuronal death in dose and exposure time dependent manner, and decreased total cellular GSH contents. The patterns of the neuronal death and the GSH decrements were dependent on the individual agents. DEM (200 micrometer) induced rapid and irreversible decrement of the GSH. BSO (1 mM) also decreased the GSH irreversibly but the rate of decrement was more progressive than that of DEM. Phorone (1 mM) reduced the GSH content to 40% by 4 hr exposure, that is comparable to the decrement of BSO, but the GSH recovered and reached over the control value by 36 hr exposure. BSO showed a minimal neurotoxicity (0-10%) at the end of 24 hr exposure, but marked neuronal cell death at the end of 48 hr exposure. The BSO (1 mM)-induced neurotoxicity was markedly inhibited by trolox or CHX and partially attenuated by MK-801. DEM induced dose-dependent cytotoxicity at the end of 24 hr exposure. Over the doses of 400 micrometer, glial toxicity also appeared. DEM (200 micrometer)-induced neurotoxicity was markedly inhibited by trolox or PDTC. Phorone (1 mM) induced moderate neurotoxicity (40%) at the end of 48 hr exposure. Only CHX showed significant inhibitory effect on the phorone-induced neurotoxicity. These results suggest that the GSH depletors induce neuronal injury via different mechanisms and that GSH depletors should be carefully employed in the researches of neuronal oxidative injuries.
Animals ; Buthionine Sulfoximine ; Cell Culture Techniques* ; Cell Death ; Cycloheximide ; Dizocilpine Maleate ; Glutathione* ; Mice* ; Neurons

OBJECTIVES: We attempted to compartmentalize the periaqueductal gray (PAG) of the rabbit in terms of the different distribution patterns between NADPH-diaphorase (NADPHd)- and calbindin D28K (CB)-positive neurons. METHODS: Immunohistochemistry and immunofluorescent labelling for CB and histochemistry for NADPHd were carried out on coronally-sectioned midbrain slices of the rabbit. RESULTS: NADPHd-positive neurons were selectively localized in the dorsolateral (DL), the middle one-third of the lateral (L), the dorsal half of the ventrolateral (VLd) PAG, and the supraoculomotor cap nucleus (Su3C). Clusters of CB-immunoreactive perikarya marked the dorsal half of DL (DLd), Su3C, the ventral one-third of L, and the ventral half of the ventrolateral (VLv) PAG. Double labelling for NADPHd and CB revealed that two markers labelled different neuronal groups in DLd and Su3C subdivisions. CONCLUSION: The present data suggest that NADPHd and CB can be regarded as reliable neurochemical markers to reveal the longitudinally-columnar organization within the PAG and to subdivide each columnar area.
Calcium-Binding Protein, Vitamin D-Dependent ; Immunohistochemistry ; Mesencephalon ; Neurons ; Periaqueductal Gray