Shu Kuei Huang

Background: Regulation of glutamate release is crucial for maintaining normal brain function, but excess glutamate release is implicated in many neuropathological conditions. Therefore, the minimum glutamate release from presynaptic nerve terminals is an important neuroprotective mechanism. Objective: In this mini-review, we analyze the three B vitamins, namely vitamin B2 (riboflavin), vitamin B6 (pyridoxine), and vitamin B12 (cyanocobalamin), that affect the 4-aminopyridine (4- AP)-evoked glutamate release from presynaptic nerve terminal in rat and discuss their neuroprotective role. Methods: In this study, the measurements include glutamate release, DiSC3(5), and Fura-2. Results: The riboflavin, pyridoxine, and cyanocobalamin produced significant inhibitory effects on 4-aminopyridine-evoked glutamate release from rat cerebrocortical nerve terminals (synaptosomes) in a dose-dependent relationship. These presynaptic inhibitory actions of glutamate release are attributed to inhibition of physiologic Ca2+-dependent vesicular exocytosis but not Ca2+-independent nonvesicular release. These effects also did not affect membrane excitability, while diminished cytosolic (Ca2+)c through a reduction of direct Ca2+ influx via Cav2.2 (N-type) and Cav2.1 (P/Q-type) Ca2+ channels, rather than through indirect Ca2+ induced Ca2+ release from ryanodine-sensitive intracellular stores. Furthermore, their effects were attenuated by GF109203X and Ro318220, two protein kinase C (PKC) inhibitors, suggesting suppression of PKC activity. Taken together, these results suggest that riboflavin, pyridoxine, and cyanocobalamin inhibit presynaptic vesicular glutamate release from rat cerebrocortical synaptosomes, through the depression Ca2+ influx via voltage- dependent Cav2.2 (N-type) and Cav2.1 (P/Q-type) Ca2+ channels, and PKC signaling cascade. Conclusion: Therefore, these B vitamins may reduce the strength of glutamatergic synaptic transmission and is of considerable importance as potential targets for therapeutic agents in glutamate- induced excitation-related diseases.

Background: Breastfeeding is an important health concern for postpartum women. Objective: This study aimed to investigate the effect of breastfeeding frequency on the level of serum prolactin (PRL), milk intake, and infant weight gain. Materials and Methods: The time and duration of each breastfeeding episode were recorded by participants from day 1 to 28 postpartum. According to their diaries, we divided participants into the low-frequency breastfeeding group (Group I; / = 10 breastfeeding episodes/day). A total of 23 mother–infant pairs were enrolled; blood samples were drawn between 1600 and 1800 hours. The PRL levels were examined using the DPC Immulite system. Results: Overall, 71.8% (23) of the enrolled mother–infant pairs completed the follow-up. Infant birth weight was higher in Group II than in Group I (3275.6 ± 93.3 g versus 2918 ± 82.1 g). On day 28 postpartum, infants in Group II ingested significantly more milk per feeding (71.6 ± 4.0 mL versus 54.1 ± 5.2 mL) and gained more weight from birth (142.9% ± 4.5% versus 130.2% ± 2.4%) compared with those in Group I. The mothers of Group II had significantly higher basal serum PRL levels (116.4 ± 11.8 ng/mL versus 72.7 ± 7.77 ng/mL), but a significantly lower increase in PRL postsuckling (168.5% ± 23.1% versus 291.6% ± 37.6% of basal PRL). The frequency of suckling was positively correlated ( r = 0.5) with the basal PRL level. Moreover, infant weight gain was significantly higher in male (144.7% ± 4.7%) than in female (132.3% ± 2.9%) infants. Conclusions: Increase in frequency of breastfeeding of over 10/day is associated with baseline PRL levels and increased milk production and weight gain. These results provide useful information for breastfeeding women.

The effect of cycloheterophyllin, a prenylflavone isolated from Artocarpus heteophyllus, on glutamate release was studied in the rat hippocampus using synaptosome and slice preparations. In rat hippocampal synaptosomes, cycloheterophyllin inhibited 4-aminopyridine (4-AP)-evoked glutamate release and elevation of intrasynaptosomal calcium levels. The inhibitory effect of cycloheterophyllin on 4-AP-evoked glutamate release was prevented in the presence of the vesicular transporter inhibitor, the N and P/Q-type calcium channel blocker, and the protein kinase C (PKC) inhibitor, but was insensitive to the intracellular Ca2+-release inhibitors, protein kinase A (PKA) inhibitor and mitogen-activated/extracellular signal-regulated kinase (MEK) inhibitor. Western blotting data in synaptosomes also showed that cycloheterophyllin significantly decreased the phosphorylation of PKC. In addition, cycloheterophyllin decreased the frequency of spontaneous excitatory postsynaptic currents (sEPSCs) without influencing the amplitude of sEPSCs and glutamate-activated currents in hippocampal slices, supporting a presynaptic action. Together, these results suggest that cycloheterophyllin inhibits presynaptic glutamate release by suppressing N and P/Q-type calcium channel and PKC activity in rat hippocampus.

Allopregnanolone, an active metabolite of progesterone, has been reported to exhibit neuroprotective activity in several preclinical models. Considering that the excitotoxicity caused by excessive glutamate is implicated in many brain disorders, the effect of allopregnanolone on glutamate release in rat cerebrocortical nerve terminals and possible underlying mechanism were investigated. We observed that allopregnanolone inhibited 4‐aminopyridine (4‐AP)‐evoked glutamate release, and this inhibition was prevented by chelating the extracellular Ca ²⁺ ions and the vesicular transporter inhibitor. Allopregnanolone reduced the elevation of 4‐AP‐evoked intrasynaptosomal Ca ²⁺ levels, but did not affect the synaptosomal membrane potential. In the presence of N‐, P/Q‐, and R‐type channel blockers, allopregnanolone‐mediated inhibition of 4‐AP‐evoked glutamate release was markedly reduced; however, the intracellular Ca ²⁺ ‐release inhibitors did not affect the allopregnanolone effect. Furthermore, allopregnanolone‐mediated inhibition of 4‐AP‐evoked glutamate release was completely abolished in the synaptosomes pretreated with inhibitors of Ca ²⁺ /calmodulin, adenylate cyclase, and protein kinase A (PKA), namely calmidazolium, MDL12330A, and H89, respectively. Additionally, the allopregnanolone effect on evoked glutamate release was antagonized by the GABA _A receptor antagonist SR95531. Our data are the first to suggest that allopregnanolone reduce the Ca ²⁺ influx through N‐, P/Q‐, and R‐type Ca ²⁺ channels, through the activation of GABA _A receptors present on cerebrocortical nerve terminals, subsequently suppressing the Ca ²⁺ ‐calmodulin/PKA cascade and decreasing 4‐AP‐evoked glutamate release.

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Lycopene is a natural dietary carotenoid that was reported to exhibit a neuroprotective profile. Considering that excitotoxicity and cell death induced by glutamate are involved in many brain disorders, the effect of lycopene on glutamate release in rat cerebrocortical nerve terminals and the possible mechanism involved in such effect was investigated. We observed here that lycopene inhibited 4-aminopyridine (4-AP)-evoked glutamate release and intrasynaptosomal Ca²⁺ concentration elevation. The inhibitory effect of lycopene on 4-AP-evoked glutamate release was markedly reduced in the presence of the Ca_v2.2 (N-type) and Ca_v2.1 (P/Q-type) channel blocker ω-conotoxin MVIIC, but was insensitive to the intracellular Ca²⁺-release inhibitors dantrolene and CGP37157. Furthermore, in the presence of the protein kinase C inhibitors GF109203X and Go6976, the action of lycopene on evoked glutamate release was prevented. These results are the first to suggest that lycopene inhibits glutamate release from rat cortical synaptosomes by suppressing presynaptic Ca²⁺ entry and protein kinase C activity.