There is accumulating evidence to implicate the importance of N-methyl-D-aspartate (NMDA) receptors to the induction and maintenance of central sensitization during pain states. However, NMDA receptors may also mediate peripheral sensitization and visceral pain. NMDA receptors are composed of NR1, NR2 (A, B, C, and D), and NR3 (A and B) subunits, which determine the functional properties of native NMDA receptors. Among NMDA receptor subtypes, the NR2B subunit-containing receptors appear particularly important for nociception, thus leading to the possibility that NR2B-selective antagonists may be useful in the treatment of chronic pain.
Several applications of pluripotent stem cell (PSC)-derived cardiomyocytes require elimination of undifferentiated cells. A major limitation for cardiomyocyte purification is the lack of easy and specific cell marking techniques. We found that a fluorescent dye that labels mitochondria, tetramethylrhodamine methyl ester perchlorate, could be used to selectively mark embryonic and neonatal rat cardiomyocytes, as well as mouse, marmoset and human PSC-derived cardiomyocytes, and that the cells could subsequently be enriched (>99% purity) by fluorescence-activated cell sorting. Purified cardiomyocytes transplanted into testes did not induce teratoma formation. Moreover, aggregate formation of PSC-derived cardiomyocytes through homophilic cell-cell adhesion improved their survival in the immunodeficient mouse heart. Our approaches will aid in the future success of using PSC-derived cardiomyocytes for basic and clinical applications.
The actions of opioid receptor agonists on synaptic transmission in substantia gelatinosa (SG) neurones in adult (6‐ to 10‐week‐old) rat spinal cord slices were examined by use of the blind whole‐cell patch‐clamp technique. Both the μ‐receptor agonist DAMGO (1 μM) and the δ‐receptor agonist DPDPE (1 μM) reduced the amplitude of glutamatergic excitatory postsynaptic currents (EPSCs) which were monosynaptically evoked by stimulating Aδ afferent fibres. Both also decreased the frequency of miniature EPSCs without affecting their amplitude. In contrast, the κ‐receptor agonist U‐69593 (1 μM) had little effect on the evoked and miniature EPSCs. The effects of DAMGO and DPDPE were not seen in the presence of the μ‐receptor antagonist CTAP (1 μM) and the δ‐receptor antagonist naltrindole (1 μM), respectively. Neither DAMGO nor DPDPE at 1 μM affected the responses of SG neurones to bath‐applied AMPA (10 μM). Evoked and miniature inhibitory postsynaptic currents (IPSCs), mediated by either the GABAA or the glycine receptor, were unaffected by the μ‐, δ‐ and κ‐receptor agonists. Similar results were also obtained in SG neurones in young adult (3‐ to 4‐week‐old) rat spinal cord slices. These results indicate that opioids suppress excitatory but not inhibitory synaptic transmission, possibly through the activation of μ‐ and δ‐ but not κ‐receptors in adult rat spinal cord SG neurones; these actions are presynaptic in origin. Such an action of opioids may be a possible mechanism for the antinociception produced by their intrathecal administration.
SUMMARY1. The effects of hypoxia on the rat hippocampal CAI neurones in tissue slices of the rat brain were studied in vitro by intracellular recording.2. In response to superfusion of a hypoxic medium equilibrated with 95 % N2-5 % C02, a majority of the neurones showed a hyperpolarization of5-15 mV in amplitude and 4-12 min in duration. The hyperpolarization was, in turn, followed by a slow depolarization which within 20 min of hypoxic exposure reached a plateau level of about 25 mV above the pre-hypoxic resting potential. Both the initial hyperpolarization and subsequent depolarization were associated with a reduction in membrane resistance. 3. The hyperpolarization reversed in polarity at a membrane potential of -83 mV. There was an almost linear relationship between amplitude of the hyperpolarization and membrane potential. The hyperpolarization was markedly enhanced in potassium-free media and was depressed in high-potassium solutions.4. The hyperpolarization was not significantly affected by low-chloride or lowsodium medium or by solution containing tetraethylammonium (10 mM), 4-aminopyridine (1-5 mM) or caesium (3 mM). Moreover, intracellular injection of ethyleneglycol-bis-(fi-aminoethylether)N,N-tetraacetic acid (EGTA) did not alter the hyperpolarization. On the other hand, barium (0 5 mM)-containing medium reduced the amplitude of the hyperpolarization by 20-40 %.5. Superfusion of ouabain (5-7 puM)-containing medium in normoxic conditions produced hyperpolarizing and depolarizing responses similar to those elicited by hypoxic exposure. The slow depolarization was also mimicked by elevation of the extracellular potassium concentration to 10-20 mm.6. Evoked i.p.s.p.s were abolished within 4 min of hypoxic exposure while evoked e.p.s.p.s were maintained for about 20 min of hypoxic superfusion. Soma spikes of the neurones elicited by a depolarizing pulse were also well preserved. Their threshold was, however, raised, concomitant with a decrease in the peak amplitude.7. When the slice was reoxygenated after 20-40 min of hypoxic exposure, the neurones immediately began to repolarize and showed a transient hyperpolarization of 5-10 mV in amplitude and 1-2 min in duration. The membrane potential, input t To whom all correspondence and reprint requests should be sent.5-2 N. FUJIWARA AND OTHERS resistance and action potential returned to the pre-hypoxic levels after 15-20 min of reoxygenation. The amplitude of the reoxygenation-induced hyperpolarization was not significantly changed when the membrane was hyperpolarized or depolarized. The hyperpolarization was eliminated by potassium-free medium or solution containing ouabain (1 ,tM).8. In a minority of the neurones the slow depolarization was suddenly followed by a rapid depolarization, after which the neurones showed no functional recovery. Such an abrupt and irreversible depolarization appeared when the slow depolarization reached membrane potentials of -30 to -40 mV.9. The results suggest that hypoxia-induced hyperpolarization is due to an increase in voltag...
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