Macroscopic brain gray matter staining: historical protocol overview and neuroanatomy learning applications in second-year medical students.

Macroscopic staining in anatomical samples of the central nervous system is a technique that has been used for decades to achieve better differentiation of multiple gray matter structures, such as the cortex, basal ganglia, and cerebellar nuclei. Staining methods are based on using the different components of the brain, mainly the lipids present in the white matter. These techniques have been progressively forgotten while computer renderings are increasing; however, as a primary exposure to surgical anatomy, stained brain specimens are considered a helpful tool. We aim to summarize different staining techniques, their principles, and their current applications for neuroanatomy learning purposes. In total, four gray matter staining protocol descriptions (Mulligan's, Roberts's, Alston's, and Prussian Blue) were performed, as well as Likert scale surveys of second-year medical students about their perceptions of the stained sections. The results showed that the different macroscopic stains for brain tissue are based on lipid and reactant interactions, intending to increase the white matter (WM) and gray matter (GM) contrast. The search also showed that most staining protocols would take 2 days to develop. Efficient preservation options include submerging the sections in formaldehyde solutions, formaldehyde-free solutions, ethanol, or applying plastination techniques. Based on the student's perspective, the stained slices seem to be a valuable alternative to facilitate the study and identification of the basal ganglia and their relationships with the white matter (from 51.2 to 72% based on the Likert scale) compared with the non-stained sections. In conclusion, macroscopic staining of brain tissue continues to be a valuable tool for comprehensively studying the brain. Further research is needed to determine the efficacy of stained specimens as teaching tools.

Evaluating the performance of WHO and CDC case definitions for influenza-like illness in diagnosing influenza during the 2022-2023 flu season in Mexico.

OBJECTIVE: This study aimed to evaluate the performance of the World Health Organization (WHO) and Centers for Disease Control and Prevention (CDC) case definitions for influenza-like illness (ILI) in diagnosing influenza during the 2022-2023 flu season in Mexico. STUDY DESIGN: We conducted a cross-sectional analysis of national epidemiological surveillance data in Mexico, focusing on respiratory viral pathogens. METHODS: We analyzed data from 6027 non-hospitalized patients between 5 and 65 years old who underwent molecular testing for respiratory viral pathogens. The performance of both case definitions was evaluated in terms of sensitivity, specificity, and the area under the receiver operating characteristic (AUROC) curve. RESULTS: Overall, the diagnostic accuracy of the evaluated ILI definitions in identifying influenza patients was low, particularly among older patients. When compared to the CDC, the WHO definition had a lower sensitivity but a higher specificity, resulting in a higher AUROC (P = 0.031) for the WHO criteria. CONCLUSIONS: Our findings suggest that the WHO and CDC ILI case definitions have limited accuracy for diagnosing influenza in non-hospitalized patients and highlight the need for more specific diagnostic tools to improve the detection of influenza cases during the flu season.

Decreased risk of COVID-19 pneumonia in children and adolescents during the Delta variant emergence.

OBJECTIVES: This study aimed to evaluate factors associated with the risk of COVID-19 pneumonia in children (aged <10 years) and adolescents (aged 10-19 years) before (March 2020-April 2021) and during (May-July 2021) the Delta (B.1.617.2) variant emergence. STUDY DESIGN: A retrospective and nationwide cohort study was conducted in Mexico. METHODS: Data from 26,961 laboratory-confirmed cases of COVID-19 were analyzed. Risk ratios (RRs) and 95% confidence intervals (CIs) were used to evaluate the association of the evaluated exposures with the risk of COVID-19 pneumonia. RESULTS: The overall incidence rate of pneumonia was 23.0 per 10,000 person-days, and it was lower during the Delta variant emergence (30.3 vs. 9.4 person-days, p < 0.001). In multiple analysis, a decreased risk of pneumonia was observed among those cases occurring in May 2021 or later (vs. March 2020-April 2021, RR = 0.98, 95% CI 0.97-0.99) and among older patients (RRper year = 0.998, 95% CI 0.996-0.998). Other comorbidities (namely, obesity, chronic kidney disease, diabetes mellitus, immunosuppression, or malignant tumors) were associated with an increased risk of severe COVID-19 manifestations. CONCLUSIONS: Our findings suggest that during the Delta variant emergence, children and adolescent patients were at reduced risk of COVID-19 pneumonia in Mexico. Further research is needed to identify factors determining the observed scenario.

Preclinical characterization of selective phosphodiesterase 10A inhibitors: a new therapeutic approach to the treatment of schizophrenia.

We have recently proposed the hypothesis that inhibition of the cyclic nucleotide phosphodiesterase (PDE) 10A may represent a new pharmacological approach to the treatment of schizophrenia (Curr Opin Invest Drug 8:54-59, 2007). PDE10A is highly expressed in the medium spiny neurons of the mammalian striatum (Brain Res 985:113-126, 2003; J Histochem Cytochem 54:1205-1213, 2006; Neuroscience 139:597-607, 2006), where the enzyme is hypothesized to regulate both cAMP and cGMP signaling cascades to impact early signal processing in the corticostriatothalamic circuit (Neuropharmacology 51:374-385, 2006; Neuropharmacology 51:386-396, 2006). Our current understanding of the physiological role of PDE10A and the therapeutic utility of PDE10A inhibitors derives in part from studies with papaverine, the only pharmacological tool for this target extensively profiled to date. However, this agent has significant limitations in this regard, namely, relatively poor potency and selectivity and a very short exposure half-life after systemic administration. In the present report, we describe the discovery of a new class of PDE10A inhibitors exemplified by TP-10 (2-{4-[-pyridin-4-yl-1-(2,2,2-trifluoro-ethyl)-1H-pyrazol-3-yl]-phenoxymethyl}-quinoline succinic acid), an agent with greatly improved potency, selectivity, and pharmaceutical properties. These new pharmacological tools enabled studies that provide further evidence that inhibition of PDE10A represents an important new target for the treatment of schizophrenia and related disorders of basal ganglia function.

Systematic study of the flow behaviour and mechanical properties of Carbopol Ultrez 10 hydroalcoholic gels.

Flow behaviour and mechanical properties of 0:100, 15:85, and 30:70 v/v gelled ethanol:water mixtures are studied as a function of pH (4.0-7.0) and Carbopol Ultrez 10 concentration (0.1-0.5%). As previously reported individually for 30% v/v alcoholic Ultrez 10 gels, flow curves were adjusted to the Ostwald's model, and similar sigmoidal dose response functions were obtained to describe the pH dependence of consistency index and mechanical properties of the systems. The concentration dependence of flow indexes was also best adjusted to one-phase exponential decay functions. As a result, the influence of ethanol content on polymer network is meaningfully assessed by means of the obtained empirical parameters: bottom value of each variable (Y(max)), pH value required for a 50% polymer network development (pH(50)), and asymptotic flow index value for the fully structured gels (n(min)). Also, it is assessed the influence of cosolvent on the above-mentioned empirical variables, by studying pH-dependence of gelation in methanol:water, 1-propanol:water and 1-butanol:water 15:85% v/v mixtures. As a result, the alcohol induced variations in consistency and mechanical properties of hydroalcoholic gels were well correlated to modifications in the solubility parameter (delta(T)).

Antisporozoite antibodies with protective and nonprotective activities: in vitro and in vivo correlations using Plasmodium gallinaceum, an avian model.

A correlation was observed between in vivo and in vitro activity of six monoclonal antibodies (mAb) against the major circumsporozoite protein of the avian malaria Plasmodium gallinaceum as follows. (1) Two mAb were protective, totally abrogating sporozoite infectivity to chicks, its natural host, in vivo; they caused 100% inhibition of sporozoite invasion (ISI) in vitro to SL-29 chicken fibroblasts and intense ISI to cultured chicken macrophages, as well as inhibited the exoerythrocytic development of sporozoites taken up by macrophages, the initial cell host of P. gallinaceum sporozoites. (2) Two mAb were partially protective in that they reduced sporozoite infectivity to chicks, caused partial ISI to SL-29 and macrophage cells and partial inhibition to the exoerythrocytic development of sporozoites in macrophages in vitro. (3) Two mAb were totally inactive in vivo although they both bound to the sporozoite antigens as detected by indirect immunofluorescence, western blot, and ELISA; they both failed to induce ISI or inhibit the exoerythrocytic development in macrophages. The possible participation of macrophages as the initial cell type involved in sporozoite destruction in the presence of anti-circumsporozoite antibodies is discussed.

The positive feedback action of vasopressin on its own release from rat septal tissue in vitro is receptor-mediated.

The effect of arginine vasopressin (AVP) on its own septal release was evaluated using an in vitro superfusion procedure. As compared to basal release from septal fragments, pulses of synthetic AVP (15 pg/5 min) resulted in a 25-fold augmented release of endogenous AVP, indicating a positive feedback action. Both the basal and stimulated AVP release were significantly increased by 60 mM potassium and markedly reduced by omission of calcium. Preincubation of the septal fragments with the V2/V1 AVP receptor antagonist d(CH2)5 [D-Tyr (Et)2,Val4]AVP resulted in a dose-dependent inhibition of the positive feedback action of AVP which was nearly completely blocked at doses between 1.25 and 5 ng per 100 microliters incubation medium. As compared to this effect, the V1 antagonist d(CH2)5 Tyr (Me)2 AVP as well as oxytocin were significantly less potent. The results suggest that the positive feedback action of AVP on its own release from septal fragments is potassium-stimulated, calcium-dependent and mainly V2 receptor-mediated. The physiological significance of this phenomenon remains to be shown.

In vitro development of exoerythrocytic forms of Plasmodium gallinaceum sporozoites in avian macrophages.

Exoerythrocytic forms of Plasmodium gallinaceum were cultured in vitro using salivary gland sporozoites extracted from experimentally infected Aedes fluviatilis mosquitoes. The host cells were macrophage precursors from chicken bone marrow. At various times after introduction of sporozoites, the cultures were stained by Giemsa or by immunofluorescence assay (IFA) using anti-sporozoite-specific monoclonal antibodies (MAb). The time to complete parasite development in vitro was 50-70 h. By 70 h, ruptured segmenters and free merozoites were visible within the cells. Inoculation of normal chickens with infected cultures induced parasitemia after a pre-patent period of 10-11 days. In vitro young exoerythrocytic forms, late schizonts that include the matured segmenters, and free merozoites shared common antigens with the sporozoites as revealed by IFA using anti-sporozoite-specific MAbs. Our data indicate that macrophages support development of P. gallinaceum sporozoites and that the circumsporozoite proteins are present until the end of the primary exoerythrocytic schizogony.

Release of vasopressin from the septum and hippocampus of freely behaving male rats: effect of bacitracin and synthetic vasopressin.

Abstract This paper describes the release characteristics of arginine vasopressin (AVP) from the dorsal septum and dorsal hippocampus of freely behaving male rats using miniaturized push-pull cannulae and a slow rate of perfusion to minimize tissue damage. The major findings were that) The spontaneous release of immunoreactive AVP (l-AVP) from the dorsal hippocampus was episodic in nature and did not require the presence of bacitracin in the perfusion medium to reliably demonstrate its pattern of release; 2) despite the fact that the septum has close to seven times more I-AVP per/mg of tissue, the dorsal septum perfusates had undetectable levels of the neuropeptide; 3) under these conditions hypertonic saline intraperitoneally did not alter the spontaneous release of this peptide; and 4) only when the perfusion medium contained effective concentrations of bacitracin (10(-4) M) were reliable and detectable levels of I-AVP measured in perfusates from the dorsal septum and exogenous synthetic AVP had a robust positive feedback action on its own release. The physiological relevance and importance of this robust and novel effect of AVP remains to be elucidated.

Positive Feedback of Vasopressin on its Own Release in the Central Nervous System: in vitro Studies.

Abstract In this paper evidence is shown that synthetic arginine vasopressin (AVP) can evoke marked in vitro release of endogenous immunoreactive AVP (I-AVP) from male rat septal and hypothalamic tissue superfused in vitro. The stimulatory action was dosedependent with a maximal amplification factor of 2.3 when using 14 pg of synthetic AVP as the stimulus. It was highly specific since only AVP was effective and not three closely related substances such as lysine vasopressin, oxytocin and a 4-9 C fragment of AVP. This reproducible effect of AVP required, however, effective concentrations of bacitracin (10(-4) to 10(-5) M) in the superfusion medium to inhibit aminopeptidase(s) capable of inactivating AVP. Lastly, the stimulatory action of AVP on its own release was not blocked by a V(1)-receptor antagonist of AVP but was blocked by a V(2)-antagonist. It is proposed that this novel and robust positive feedback of AVP on its own release may be involved in the mechanism of memory consolidation of certain behavioral tasks known to be affected by AVP.

The nature and magnitude of in vivo 5-hydroxyindoleacetic acid output from 5-hydroxytryptamine terminals is related to specific regions of the suprachiasmatic nucleus.

Eight cycling female rats were implanted with push-pull cannulae over the region of the suprachiasmatic nuclei (SCN) and allowed 7-10 days for recovery. Perfusion of the SCN continued in these freely behaving rats for 5-6 h of the light period and the subjective scotophase. The release of 5-hydroxyindoleacetic acid (5-HIAA) ranged from 10 to 350 pg 5-HIAA/min. Significantly, the amplitude and characteristics of the output of 5-HIAA were highly location dependent in that rostral cannulae placements revealed high amplitude changes with initial mean values of 40 pg 5-HIAA/min, which increased toward the dark phase to mean peak values of 195 pg 5-HIAA/min. Caudal cannulae placements revealed a low amplitude, high frequency, basal type of 5-HIAA release which did not increase toward the dark period (approximately 5 pg/min). 5-Hydroxytryptophan infusion resulted in a significant marked increase in the basal release of 5-HIAA confirming the biochemical viability of the area undergoing perfusion. These results suggest that the in vivo measurement of 5-HIAA from 5-hydroxytryptamine (5-HT) terminals in the region of the SCN could reflect discrete functional activity of serotonergic terminals within specific regions of the SCN in a freely behaving rat. Furthermore, the biochemical viability of these 5-HT terminals and the ability of the rat's SCN to exhibit marked differential changes in 5-HT activity emphasizes the physiological relevance of this model system to study neuroendocrine events in freely behaving animals.

Functional characteristics of the luteinizing hormone-releasing hormone pulse generator in conscious unrestrained female rabbits: activation by norepinephrine.

We probed the activity of the neural LHRH apparatus by means of push-pull cannulae implanted in the hypothalamus of conscious intact anestrous unrestrained New Zealand White female rabbits raised under natural light. The data revealed that the neuropeptide LHRH is released in a pulsatile manner (one pulse per 46.4 +/- 3.9 min; amplitude, 1.63 +/- 0.39 pg; n = 7) from a rather extensive area within the basal hypothalamus of the rabbit covering the rostral, medial, and posterior regions in a rostro-caudal direction and about 1 mm lateral. A remarkable change in mean LHRH output was observed during the year, and two distinct types of LHRH output were detected: low (0.72 +/- 0.07 pg/10 min; n = 7) when these reflex ovulators were perfused during late spring to early summer and high (2.92 +/- 0.29 pg/10 min; n = 5) when perfused during late summer, early fall to spring months. This 4-fold annual change in LHRH output was mainly due to changes in the amplitude of the LHRH signal and was not correlated with the position of the push-pull cannula within a restricted area in the hypothalamus. These changes may be related to the state of the ovaries, since rabbits with high LHRH output showed larger ovaries with well developed follicles than rabbits with low LHRH output. Norepinephrine, a hypothalamic neurotransmitter known to be involved in the mounting-evoked ovulation reflex in this induced ovulator, elicited a 2-fold greater mean LHRH release (P less than 0.05) when infused in doses of 10(-7) - 10(-6) M directly into the hypothalamic perfusion area. The data indicate that the activity of the LHRH pulse generator in this species presents remarkable annual variations in its mean LHRH output mainly due to a greater amplitude of the LHRH signal and that norepinephrine is a potent stimulator of LHRH release in conscious unrestrained female rabbits.

Push-pull perfusion of the hypothalamus and the caudate nucleus in conscious, unrestrained animals.

The present results further confirm the usefulness of the push-pull cannula to study the in vivo activity of several neural systems in conscious, freely moving animals. However, it is evident that the inflammatory reaction to the local injury induced by the cannula may have different consequences in the function of a particular brain area. In the caudate nucleus, the responsiveness of the dopaminergic terminals of the nigrostriatal DA system to local infusion of AMPH was clearly reduced after the first two to three weeks post cannula implant. In contrast, in the hypothalamus of the rat and the rabbit, the spontaneous fluctuating activity of the LHRH pulse generator remained unaffected during a two to three month period when several perfusions could be performed in the same animal. Further studies will be required to establish the reasons for such marked differences in response to injury of these two brain structures. Various neurochemicals can be measured simultaneously from the hypothalamus of conscious rabbits. The present results indicate that NE hypothalamic impulse flow increases during the afternoon whereas the impulse flow of DA (as estimated by DOPAC output) decreases or does not change. In rabbit No. 2, simultaneous measurement of hypothalamic NE release and LHRH release indicates an absence of correlation. In sharp contrast, the simultaneous measurement of prostaglandin E2 release and LHRH release from the hypothalamus of rabbit 2-ER revealed a closer association between these two neuronal events. Apparently the push-pull technique can be used to examine long-term seasonal as well as circadian (extended 24 hr perfusion) spontaneous changes in the activity of the LHRH pulse generator in individual animals. It seems that seasonal changes characterize the function of the rabbit LHRH pulse generator. Lastly, physiological doses of progesterone infused for short times directly into the hypothalamus of female rabbit stimulate the LHRH pulse generator of this species with marked increases in the amplitude of the LHRH signal. In conclusion, the PPP technique coupled to HPLC-EC and RIA procedures to measure localized release of neurotransmitters, metabolites, and neuropeptides simultaneously in the same perfusate samples from discrete brain areas appears to be a powerful technique in furthering our knowledge of the in vivo local neurochemistry of the brain of conscious, freely moving animals.