The effects of caffeine in the social interaction test and on exploration in rats: comparison with ethanol and clorazepate

The effects of caffeine (20mg/kg) in the holeboard and social interaction tests were compared with those of ethanol (0.4g and dipotassium clorazepate (3mg/kg), following acute administration in one group of rats or after five daily injections in another group. The rats were put in pairs into an unfamiliar arena with high levels of illumination (n = 80), or tested individually in the holeboard (n = 80). Acute caffeine produced no effect on the time spent in social interaction, although it enhanced the number of social contacts, and both genital and total sniffing. Following five injections, caffeine also increased the time spent in social interaction. Acute clorazepate enhanced this time but this effect showed partial tolerance after five injections. Clorazepate also enhanced the number and duration of social contacts, increasing social grooming and genital sniffing, regardless of the duration of the treatment. Ethanol increased the time spent in social interaction following five injections, and increased social grooming. In the holeboard, stimulant effects were observed for caffeine and clorazepate, showing partial tolerance and without any effect on head dipping. In the social interaction test, only a stimulant effect for caffeine was obtained. The results of this study suggest that, under some circumstances, caffeine may enhance social interaction, in a manner similar to standard anxiolytics. Such an effect is potentiated by repeated administration.     

Effects of capsaicin,tachykinins, calcitonin gene-related peptide and bradykinin in the pig iris sphincter muscle

Summary Capsaicin-sensitive sensory neurons of the rabbit iris, by releasing tachykinins, exert a major role in the control of pupil motility in response to various noxious stimuli. However, the contribution of sensory innervation to the regulation of iris smooth muscle tone in other mammals species is not known. We have studied the effects produced by electrical field stimulation, capsaicin, substance P, neurokinin A, calcitonin gene-related peptide (CGRP), and bradykinin in the isolated iris sphincter muscle of the pig.Capsaicin (10 M): a) contracted the isolated sphincter muscle and; b) released immunoreactivity for substance P (SP-LI) and CGRP (CGRP-LI) from this preparation. These two effects were no longer observed at the second exposure to the drug. Electrical field stimulation (10 Hz, 60 V, 0.5 ms for 5 s) produced a biphasic contractile response. The rapid component was inhibited by atropine (1 M), while the delayed response was blocked by previous exposure to capsaicin (10 M).Substance P and neurokinin A consistently produced contraction of the pig iris sphincter muscle, substance P being more potent than neurokinin A. CGRP induced a contractile response in more than 50% of the preparations. The tachykinin antagonist [D-Argl, D-Trp^7,9, Leu¹¹-substance P (3 M) blocked: a) the effect of substance P (1 nM); b) the delayed response to electrical field stimulation and; c) reduced by more than 50% response to capsaicin. Bradykinin (10 M) failed to release either SP-LI or CGRP-LI. The contractile response evoked by bradykinin was unaffected by in vitro pretreatment with capsaicin (10 M).The existence in the pig iris of capsaicin-sensitive sensory fibres releasing neuropeptides and thus regulating sphincter muscle tone is proposed. Send offprint requests to Dr. P. Geppetti at the above address     

Current immunoassays and detection of antibodies elicited by Omicron SARS-CoV-2 infection

The present study aimed to determine whether current commercial immunoassays are adequate for detecting anti-Omicron antibodies. We analyzed the anti-SARS-CoV-2 antibody response of 23 unvaccinated individuals 1–2 months after an Omicron infection. All blood samples were tested with a live virus neutralization assay using a clinical Omicron BA.1 strain and four commercial SARS-CoV-2 immunoassays. We assessed three anti-Spike immunoassays (SARS-CoV-2 IgG II Quant [Abbott S], Wantaï anti-SARS-CoV-2 antibody ELISA [Wantaï], Elecsys Anti-SARS-CoV-2 S assay [Roche]) and one anti-Nucleocapsid immunoassay (Abbott SARS-CoV-2 IgG assay [Abbott N]). Omicron neutralizing antibodies were detected in all samples with the live virus neutralization assay. The detection rate of the Abbott S, Wantai, Roche, and Abbott N immunoassays were 65.2%, 69.6%, 86.9%, and 91.3%, respectively. The sensitivities of Abbott S and Wantai immunoassays were significantly lower than that of the live virus neutralization assay (p = 0.004, p = 0.009; Fisher's exact test). Antibody concentrations obtained with anti-S immunoassays were correlated with Omicron neutralizing antibody concentrations. These data provide clinical evidence of the loss of performance of some commercial immunoassays to detect antibodies elicited by Omicron infections. It highlights the need to optimize these assays by adapting antigens to the circulating SARS-CoV-2 strains.     

Positive SARS-CoV-2 RT-qPCR of a nasal swab spot after 30 days of conservation on filter paper at room temperature

We tested the use of nasal swabs spotted onto filter paper (Whatman 3M) for the molecular diagnosis of SARS-CoV-2 infection. Spots of a positive nasal swab in conservation medium (B.1.177 strain, 21Ct) were still positive (duo E-gene/IP4) after 10, 20, and 30 days of conservation at room temperature, with Ct values of 28, 27, and 26, respectively. Direct spotting of the swab at bedside (omicron strain) still gave a positive result after 10 days in two RT-qPCR systems: 33.7 Ct using duo E-gene/IP4, and 34.8 using a specific Omicron system. Spotting of a dilution range of media spiked with the Delta (strain 2021/FR/0610, lineage B 1.617.2) and Omicron strains (strain UVE/SARS-CoV-2/2021/FR/1514) showed a threshold of 0.04 TCID₅₀ after 10 days of conservation. We show, for the first time, that this simple and low-cost conservation method can be used to store samples for RT-qPCR against SARS-CoV-2 for up to at least 1 month.     

Prevalence and control of arterial hypertension in the south-east of Spain: A radical but still insufficient improvement

Stroke mortality rates in Spain are one of the highest in all of Europe. At the same time, the Murcia region (south-east Spain) shows, for both genders, the highest age-adjusted stroke mortality rates in all of Spain. The earliest available hypertension figure estimations for this area go back to 1981, when a high prevalence combined with an almost nonexistent control was detected. One decade later, updated prevalence estimations of hypertension are presented jointly with their degree of control and their association with other risk factors based on the results of a prevalence survey in a random population sample (n = 3,091). Arterial blood pressure was measured following the MONICA protocol, maintaining a tight quality control on between and within-observer variability. As hypertensive was considered as any person with systolic blood pressure (SBP) 140 mmHg or with diastolic blood pressure (DBP) 90 mmHg or with antihypertensive pharmacological treatment. Detected prevalence rises to 32.3% (CI 95%: 29.1%–35.5%) among males and to 23.7% (CI 95%: 21.4%– 26%) among females, maintaining its level regarding figures observed back in 1981. However, its control has been increased, especially among hypertensive women [from less than 5% in 1981 to 35% (95% CI: 32%–37.8%) at present]. Hypertension is strongly associated to hipercholesterolemia, overweight, obesity and diabetes (p < 0.01). On the contrary, it shows an opposite association with current smoking, higher educational level and leisure time physical activity (p < 0.01). The highest educational level was associated with better hypertension treatment and control. In summary, while hypertension prevalence is stabilized in our population, its control has improved in a measurable but still insufficient way. These results are in accordance with a decreasing trend in stroke mortality registered in the Murcia Region along the last decade.     

Reduction of urinary 8-epi-prostaglandin F2α during cyclo-oxygenase inhibition in rats but not in man

1. 8-epi-prostaglandin (PG) F_2α, a major F₂ isoprostane, is produced in vivo by free radical-dependent peroxidation of lipid-esterified arachidonic acid. Both cyclo-oxygenase isoforms (COX-1 and COX-2) may also form free 8-epi-PGF_2α as a minor product. It has been recently seen in human volunteers that the overall basal formation of 8-epi-PGF_2α in vivo is mostly COX-independent and urinary 8-epi-PGF_2α is therefore an accurate marker of ‘basal'' oxidative stress in vivo.
2. To test the validity of this marker in the rat, we evaluated in vivo the effect of COX inhibition on the formation of 8-epi-PGF_2α vs prostanoids. Two structurally unrelated COX inhibitors (naproxen: 30 mg kg⁻¹ day⁻¹; indomethacin: 4 mg kg⁻¹ day⁻¹) were given i.p. to rats kept in metabolic cages. In vivo formation of 8-epi-PGF_2α was assessed by measuring its urinary excretion. Prostanoid biosynthesis was assessed by measuring urinary excretion of major metabolites of thromboxane (TX) and prostacyclin (2,3-dinor-TXB₁ and 2,3-dinor-6-keto-PGF_1α). All compounds were selectively measured by immunopurification/gas chromatography-mass spectrometry.
3. Naproxen reduced urinary excretion of 2,3-dinor-TXB₁ and 2,3-dinor-6-keto-PGF_1α but, unexpectedly, also that of 8-epi-PGF_2α (82, 49 and 52% inhibition, respectively). Indomethacin had a similar effect (77, 69 and 55% inhibition). Esterified 8-epi-PGF_2α in liver and plasma remained unchanged after indomethacin.
4. These findings prompted us to re-assess the contribution of COX activity to the systemic production of 8-epi-PGF_2α in man. We gave naproxen (1 g day⁻¹) to healthy subjects (four nonsmokers and four smokers). Urinary 8-epi-PGF_2α remained unchanged in the two groups (9.63±0.99 before vs 10.24±1.01 after and 20.14±3.00 vs 19.03±2.45 ng h⁻¹ 1.73 m⁻²), whereas there was a marked reduction of major urinary metabolites of thromboxane and prostacyclin (about 90% for both 11-dehydro-TXB₂ and 2,3-dinor-TXB₂; >50% for 2,3-dinor-6-keto-PGF_1α).
5. To investigate whether rat COX-1 produces 8-epi-PGF_2α more efficiently than human COX-1, we measured the ex vivo formation of 8-epi-PGF_2α and TXB₂ simultaneously in whole clotting blood. Serum levels of 8-epi-PGF_2α and TXB₂ were similar in rats and man.
6. We conclude that a significant amount of COX-dependent 8-epi-PGF_2α is present in rat but not in human urine under normal conditions. This implies that urinary 8-epi-PGF_2α cannot be used as an index of near-basal oxidant stress in rats. On the other hand, our data further confirm the validity of this marker in man.

     

Evidence for the involvement of the nitric oxide–cGMP pathway in the antinociception of morphine in the formalin test

The effect of inhibition of nitric oxide synthesis and guanylate cyclase on the peripheral antinociceptive effect of morphine was assessed by using the formalin test in the rat. Saline, N^G-monomethyl-

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-arginine, a nitric oxide synthesis inhibitor (50 μg) and methylene blue, a guanylate cyclase inhibitor (500 μg), did not exhibit any antinociceptive activity. However, morphine (10 μg) produced a significant antinociceptive effect in phases 2a and 2b, which was reduced by pretreatment with either N^G-monomethyl-

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-arginine or methylene blue. These results suggest that the local administration of morphine induces antinociception by the activation of the

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-arginine–nitric oxide–cGMP pathway.     

Differential effects of zidovudine and zidovudine triphosphate on mitochondrial permeability transition and oxidative phosphorylation

1. The effects of zidovudine (ZDV) and zidovudine triphosphate (ZDV-3P) on Ca²⁺-induced mitochondrial permeability transition (MPT), respiratory control ratio (RCR) and ATP synthesis have been investigated on isolated rat liver mitochondria.
2. ZDV slightly but significantly decreased RCR and ATP synthesis but was ineffective in inhibiting MPT. In contrast, ZDV-3P did not alter RCR and ATP synthesis but strongly inhibited MPT (IC₅₀=3.0±0.9 μM).
3. The effect of ZDV-3P on mitochondrial swelling required a preincubation time. When incubated 10 min with mitochondria, ZDV-3P (8 μM) totally inhibited the rate of swelling.
4. ADP, ATP and atractyloside, which are agents known to interact with the mitochondrial adenine nucleotide carrier (ANC), antagonized the effect of ZDV-3P on mitochondrial swelling. Indeed, the IC₅₀ value of ZDV-3P increased from 3.0 to 17.4, 93.6 and 66.5 μM, in the presence of 20 μM, ADP, ATP or atractyloside, respectively.
5. ZDV-3P did not displace [³H]-ATP from its mitochondrial binding site(s) whereas ADP and atractyloside did, suggesting that ZDV-3P and [³H]-ATP do not share the same binding sites.
6. ZDV-3P did not affect either mitochondrial respiration or ATP synthesis but inhibited Ca²⁺-dependent mitochondrial swelling. It was concluded that mitochondrial toxic effects observed during the chronic administration of ZDV cannot be related to its active metabolite (ZDV-3P).