Tomoko Kadota

Essential role of D1 but not D2 receptors in methamphetamine‐induced impairment of long‐term potentiation in hippocampal–prefrontal cortex pathway

Methamphetamine (MA) abuse induces deficits in cognitive performance that are related to dysfunction of the prefrontal cortex (PFC). The medial portion of the prefrontal cortex (mPFC) in rats that is crucial for cognitive function has been shown to undergo long‐term potentiation (LTP) in the projections from the hippocampus. However, no study has been performed to evaluate the influence of MA on synaptic plasticity in the hippocampal–mPFC pathways. In the present experiments, we investigated the effects of repeated MA administration on hippocampal–mPFC LTP, together with MA‐induced stereotyped behaviors. Repeated MA administration produced behavioral sensitization and LTP impairment in the hippocampal–mPFC pathways. The MA‐induced impairment of hippocampal–mPFC LTP was prevented by the pretreatment of dopamine 1 (D1) but not dopamine 2 (D2) receptor antagonists, while D1 and D2 receptor antagonists attenuated the MA‐induced stereotyped behaviors. These findings suggest that D1 receptors are crucial for the MA‐induced deterioration of synaptic plasticity in the hippocampal–mPFC circuits. Impairment of LTP associated with D1 receptor dysfunction may underlie cognitive deficits in MA‐dependent subjects.

Rapid structural remodeling of shaft synapses associated with long-term potentiation in the cat superior cervical ganglion in situ.

Synaptic plasticity associated with long-term potentiation was studied electrophysiologically and ultrastructurally in the cat superior cervical ganglion in situ. The preganglionic nerve fiber was stimulated at 10 Hz for 50 s for conditioning and then at 1 Hz for 1-3 h to monitor changes in the postganglionic compound action potential (PGP). The present material has shown the long-term potentiation (LTP), around 120% of the control, which lasted for up to 3 h. Fifteen of 18 ganglia (83%) have shown LTP. Ultrastructural studies demonstrated the synaptic structural remodeling: (1) The preganglionic nerve terminals ordinarily made mainly asymmetrical type of shaft synapses directly with dendrites of the ganglion cells that lacked dendritic spines; (2) conditioning tetanus rapidly remodeled simple shaft synapses into perforated ones characterized by perforations in the postsynaptic density (PSD), some of which had synaptic spinules associated with the perforated PSDs, i.e. spinule-synapses; (3) a rapid increase in the number of both structures was detected immediately after the tetanus. Perforated synapses and the spinule-synapses increased from 5% and 0% in the control to 27 and 9% at 0 min, respectively. Spinule-synapses occurred about one-third of the perforated shaft synapses; (4) Increased numbers of restructured shaft synapses was maintained for 15 min in ganglia expressing LTP; (5) Remodeled synapses did not increase in ganglia that did not express LTP or ganglia that were activated at 0.5 or 1 Hz. It was suggested a rapid increase in the number of remodeled synapses associated with the onset of LTP and its durability at its earlier phases in the cat SCG.

Dopamine D1 and D2 Receptors and Their Signal System Present in Coated Vesicles Prepared from Bovine Striatal Tissue

Abstract: Coated vesicles (CVs) isolated from bovine striatal tissue were examined to determine whether they are associated with dopamine signal systems consisting of dopamine D1 and D2 receptors, G proteins, and adenylate cyclase. Dopamine receptors in CVs were characterized by a dopamine D1 receptor antagonist, [3H]SCH 23390, and a dopamine D2 receptor antagonist, [3H]‐spiroperidol. The bindings of both ligands were specifically saturable and reversible with a dissociation constant (KD) of 0.65 and 0.5 nM, respectively. Dopaminergic antagonists and agonists inhibited the specific bindings of [3H]SCH 23390 and [3H]spiroperidol in a stereoselective and concentration‐dependent manner with an appropriate rank order potency for dopamine D1 or D2 receptors. The regulations of the agonist binding by guanyl‐5‐ylimidodiphosphate were observed. ADP ribosylation of the CVs with [32P]NAD demonstrated predominant labeling of bands of Mr 47,000–52,000, 42,000–45,000, and 40,000‐39,000, which corresponded to the known molecular weights of the α subunits of Gs and Gi proteins. The presence of α and β subunits of G proteins in the CVs was also confirmed by immunoblotting assay. Adenylate cyclase activity, which was stimulated by SKF 38393 and inhibited by dopamine D2 receptor agonists, was present in the CVs. These findings suggest that the dopamine D1 and D2 receptors in the CVs couple with adenylate cyclase via Gs/Gi protein.

Characterization of [3H]5-hydroxytryptamine and [3H]spiperone binding sites in clathrin-coated vesicles from bovine brain

Coated vesicles prepared from bovine brain cerebral cortex exhibited [3H]5-hydroxytryptamine (5-HT, serotonin) and [3H]spiperone binding activities. The binding activities were localized in the inner core vesicles. Binding reached an equilibrium level by 30-45 min at 30 degreesC, and was reversed by the addition of 100 microM 5-HT for [3H]5-HT binding or 10 microM ketanserin for [3H]spiperone binding. The saturation binding experiments indicated a single class of binding sites for [3H]5-HT and [3H]spiperone with apparent Kd values of 2.4 and 1.75 nM, respectively. The binding of [3H]5-HT was displaced by 5-HT and 8-hydroxy-2-(di-n-propylamino)-tetralin (8-OH-DPAT), but not by ketanserin. The binding of [3H]spiperone was displaced by spiperone and ketanserin but not by 5-HT or 8-OH-DPAT even at 1 mM. The coated vesicles were shown by immunoblotting assay to contain alpha-subunits of GTP-binding proteins, Galphas, Galphai2, Galphai3, Galphao and Galphaq/11. Forskolin-stimulated adenylate cyclase activity in the coated vesicles was inhibited to 80% of the control level by 5-HT or 8-OH-DPAT. These results suggested that 5-HT1A and 5-HT2A receptors are present in bovine brain coated vesicles and that the 5-HT1A receptors are coupled to adenylate cyclase activity via GTP binding proteins.

Rapid recovery of structure and function of the cholinergic synapses in the cat superior cervical ganglionin vivo following stimulation-induced exhaustion

SummaryCat superior cervical ganglia (SCG) were tetanically stimulatedin vivo at 30–100 Hz until neural transmission was exhausted, and then were allowed to rest and recover. Changes in their cholinergic synapses were examined electrophysiologically and morphologically during the time of tetanic stimulation and during recovery. For morphometric analysis the presynaptic terminal was subdivided into two areas: an area directly over the active zone, termed zone-I, (bounded by a hemicircle with a diameter equivalent to the active zone length), and the remaining preterminal area, termed zone-II. In control ganglia before stimulation synaptic vesicle density in zone-I (SVD-I) averaged 90 μm−2 and the number of vesicles actually attached to the active zone (SVA) averaged about 2.5 per single profile of nerve terminal. Upon stimulation, the postganglionic potential immediately began to decline in amplitude and disappeared after 1 min of stimulation. Simultaneously, SVD-I declined to less than 35 μm−2 and SVA declined to less than 1 per section. Thereafter, stimulation was terminated and the ganglion was allowed to rest. Recovery of the postganglionic potential was monitored by stimulation at 1 Hz. The postganglionic potential reached control levels after only 1 min of rest. Likewise, the structural parameters, SVD-I and SVA, also rapidly recovered, reaching control levels after only 30 sec of rest, slightly faster than the postganglionic potential. This illustrates that stimulation-induced fatigue of transmitter output and depletion of synaptic vesicles recover to the control level at a high rate in synapses of the cat SCG with a normal supply of blood. In fact, morphological recovery may be slightly faster than electrophysiological recovery. Mechanisms of vesicle formation and migration to the presynaptic area are discussed in light of these observations.

Poststimulation Increase of Synaptic Vesicle Number in the Preganglionic Nerve Terminals of the Cat Sympathetic Ganglion InVivo

Vesicular release of neurotransmitter has been suggested in a variety of nerve endings by morphological reports in which tetanic stimulation is shown to cause the reduction of synaptic vesicles (SVs) in number and simultaneously to depress the postsynaptic response (Ceccarelli et al., 1973; Heuser and Reese, 1973; Pysh and Wiley, 1974; Zimmerman and Whittaker, 1974a, b). Vesicular release of transmitter via exocytosis is supposed to be followed by the subsequent reformation of vesicles by endocytotic retrieval of terminal plasmamembrane by coated vesicles (recycling hypothesis of synaptic-vesicle membrane) (Heuser and Reese, 1973). Questions remains, however, concerning the vesicle hypothesis of the release of quanta of neurotransmitter (Zimmerman, 1979; Ceccarelli and Hurlbut, 1980; Tauc, 1979; Israel and Manarche, 1985). In addition, recent articles have shown the low rate of coated vesicles in the SV reformation, having implied the necessities of examinations for other mechanisms for supplying SVs during transmitter release (Kadota and Kadota, 1982; Meshul and Pappas, 1984; Parducz, 1986; Torri-Tarelli et al., 1987). The purpose of this study is to examine the ultrastructural changes in an neuro-neuronal synapse under normal supply of blood. The preganglionic nerve terminal of the cat superior cervical ganglion (SCG) was employed as the experimental material in the present experiment. It was simple to maintain this ganglion under intact blood sypply and to fix rapidly by perfusion via the lingual artery (Kadota and Kadota, 1982).

Localization of a 82 kDa protein in postsynaptic density and its association with cytoskeletons

A fraction of synaptic junctional complex (SJC) was prepared from rat synaptosomes and served as antigen material to produce monoclonal antibodies (Mab) for examining the component proteins of the SJC. An antibody, Mab SJ-8, was obtained, which recognized a protein with a molecular weight of 82,000 Da in the SJC preparation by immunoblot analysis. The immunohistochemical localization of the 82 kDa protein was studied with the rat cerebellum. Mab SJ-8 labeled the peripheral areas of the Purkinje and granule cells. Small punctate areas were also stained in the molecular layer with SJ-8. Intracellular localization of the protein was examined with rat brain synaptosomes. Immunoelectron microscopy demonstrated that Mab SJ-8 strongly labeled the postsynaptic density (PSD) and also a fibrous network spreading out of it. However, the antibody did not label the pre- or post-synaptic membrane or the cleft material.