Vascular Gap Junctions Contribute to Forepaw Stimulation-Induced Vasodilation Differentially in the Pial and Penetrating Arteries in Isoflurane-Anesthetized Rats

Somatosensory stimulation causes dilation of the pial and penetrating arteries and an increase in cerebral blood flow (CBF) in the representative region of the somatosensory cortex. As an underlying mechanism for such stimulation-induced increases in CBF, cerebral artery dilation has been thought to propagate in the vascular endothelium from the parenchyma to the brain surface. Vascular gap junctions may propagate vasodilation. However, the contribution of vascular gap junctions to cerebrovascular regulation induced by somatosensory stimulation is largely unknown. The aim of the present study was to investigate the contribution of vascular gap junctions to the regulation of the pial and penetrating arteries during neuronal activity attributed to somatosensory stimulation. Experiments were performed on male Wistar rats (age: 7–10 weeks) with artificial ventilation under isoflurane anesthesia. For somatosensory stimulation, the left forepaw was electrically stimulated (1.5 mA, 0.5 ms and 10 Hz, for 5 s). The artery in the forelimb area of the right somatosensory cortex was imaged through a cranial window using a two-photon microscope and the diameter was measured. Carbenoxolone (CBX) was intravenously (i.v.) administered, at a dose of 100 mg/kg, to block vascular gap junctions. The forepaw electrical stimulation increased the diameter of the pial and penetrating arteries by 7.0% and 5.0% of the pre-stimulus diameter, respectively, without changing the arterial pressure. After CBX administration, the change in pial artery diameter during forepaw stimulation was attenuated to 3.2%. However, changes in the penetrating artery were not significantly affected. CBF was measured using a laser speckle flowmeter, together with somatosensory-evoked potential (SEP) recorded in the somatosensory cortex. The extent of CBF increase (by 24.1% of the pre-stimulus level) and amplitude of SEP were not affected by CBX administration. The present results suggest that vascular gap junctions, possibly on the endothelium, contribute to pial artery dilation during neuronal activity induced by somatosensory stimulation

A liquid crystalline phase in spermidine-condensed DNA

Over a large range of salt and spermidine concentrations, short DNA fragments precipitated by spermidine (a polyamine) sediment in a pellet from a dilute isotropic supernatant. We report here that the DNA-condensed phase consists of a cholesteric liquid crystal in equilibrium with a more concentrated phase. These results are discussed according to Flory's theory for the ordering of rigid polymers. The liquid crystal described here corresponds to an ordering in the presence of attractive interactions, in contrast with classical liquid crystalline DNA. Polyamines are often used in vitro to study the functional properties of DNA. We suggest that the existence of a liquid crystalline state in spermidine-condensed DNA is relevant to these studies

Spatial Frequency-Based Analysis of Mean Red Blood Cell Speed in Single Microvessels: Investigation of Microvascular Perfusion in Rat Cerebral Cortex

BACKGROUND: Our previous study has shown that prenatal exposure to X-ray irradiation causes cerebral hypo-perfusion during the postnatal development of central nervous system (CNS). However, the source of the hypo-perfusion and its impact on the CNS development remains unclear. The present study developed an automatic analysis method to determine the mean red blood cell (RBC) speed through single microvessels imaged with two-photon microscopy in the cerebral cortex of rats prenatally exposed to X-ray irradiation (1.5 Gy). METHODOLOGY/PRINCIPAL FINDINGS: We obtained a mean RBC speed (0.9±0.6 mm/sec) that ranged from 0.2 to 4.4 mm/sec from 121 vessels in the radiation-exposed rats, which was about 40% lower than that of normal rats that were not exposed. These results were then compared with the conventional method for monitoring microvascular perfusion using the arteriovenous transit time (AVTT) determined by tracking fluorescent markers. A significant increase in the AVTT was observed in the exposed rats (1.9±0.6 sec) as compared to the age-matched non-exposed rats (1.2±0.3 sec). The results indicate that parenchyma capillary blood velocity in the exposed rats was approximately 37% lower than in non-exposed rats. CONCLUSIONS/SIGNIFICANCE: The algorithm presented is simple and robust relative to monitoring individual RBC speeds, which is superior in terms of noise tolerance and computation time. The demonstrative results show that the method developed in this study for determining the mean RBC speed in the spatial frequency domain was consistent with the conventional transit time method

A novel techinique with quantum dot for multiphoton microscope imaging of in vivo brain vasculature

Brain Energy Metabolism and Blood Flo

Intracortical microcirculatory change induced by anesthesia in rat somatosensory cortex

International Society on Oxygen Transport to Tissue 200

Hemodynamic Response to Forepaw Stimulation in Anesthetized Mice Somatosensory Cortex

Aim: Genetically altered mice are a potentially powerful tool to elucidate the molecular basis of neuro-vascular coupling mechanism.However, neural activity-induced hemodynamic responses in anesthetized murine cortex remains uncharacterized, which hampers the use of a mice model for functional imaging studies (e.g., fMRI and optical imaging). In the present study, hemodynamic responses to forepaw stimulation in the mice somatosensory cortex were characterized under pentobarbital, ketamine-xylazine, and isoflurane anesthesia, all of which are generally used for repeated animal experiments.\nMaterials and methods: A total of thirteen male C57B6J mice (23-35 g) were divided into three groups. First group (N = 4) was induced with pentobarbital anesthesia (an initial dose of 90 mg/kg i.p. and supplemental dose of 30-45 mg/kg/h i.p.). Second group (N = 6) was induced with a cocktail of ketamine and xylazine (an initial dose of 60 mg/kg and 10 mg/kg i.m., respectively, and supplementally injected with only ketamine 20 mg/kg i.m. every 20-40 min.). The animals in the third group (N = 3) were ventilated with a mixture of isoflurane (4% for induction and 1.3-1.5% for experiments) and air with supplemental oxygen (a total of 30-35%). Animals in the isoflurane group were intubated and mechanically ventilated, whereas the other two groups breathed spontaneously. After the skin covering the skull was removed, oil was immediately placed to preserve the integrity of the skull. First, intrinsic optical imaging (620-nm wavelength) was performed for the localization of the forepaw area in the primary somatosensory cortex. Then, hemodynamic response was measured with laser-Doppler flowmetry (LDF) on the activation focus induced by electrical forepaw stimulation (rectangular pulses with 0.5-ms pulse width and 0.8-mA current). Stimulation duration was fixed at 10 sec for all experiments and stimulation rate (frequency) was varied (2, 4, 6, 8 and 12 Hz). Preliminary experiments determined a pulse width and current that would not elicit a pain response from the animal.\nResults and discussion: All animal groups showed clear localization of forepaw area in 620-nm optical imaging. The clearest contrast of activation area was observed with ketamine-xylazine group. Under pentobarbital anesthesia, the hemodynamic response increased with an increase in stimulation frequency. The peak of 10 +/- 12% (Mean +/- SD, N = 4) signal change in LDF was observed at stimulation with 12 Hz. Under ketamine-xylazine anesthesia, the highest signal change (13 +/- 4% at peak) was observed at stimulation with 6 Hz. These results indicate that the optimum frequency differs depending on the anesthesia used, which was consistent with our previous results obtained in rats (Masamoto et al., 2006). In isoflurane-anesthetized group, however, no significant hemodynamic response was observed for any stimulation frequency. This result indicates that the isoflurane may be not a good agent for functional imaging studies with mice, which was not the case in rats anesthetized with isoflurane (Masamoto et al., 2006). In conclusion, mice anesthetized with pentobarbital or ketamine-xylazine can be used for repeated functional imaging studies, which make possible to probe the mechanism of neurovascular coupling with a specific molecular basis.Brain\u2707 and BrainPET\u270

Anesthesia and the quantitative evaluation of neurovascular coupling

Anesthesia has broad actions that include changing neuronal excitability, vascular reactivity, and other baseline physiologies and eventually modifies the neurovascular coupling relationship. Here, we review the effects of anesthesia on the spatial propagation, temporal dynamics, and quantitative relationship between the neural and vascular responses to cortical stimulation. Previous studies have shown that the onset latency of evoked cerebral blood flow (CBF) changes is relatively consistent across anesthesia conditions compared with variations in the time-to-peak. This finding indicates that the mechanism of vasodilation onset is less dependent on anesthesia interference, while vasodilation dynamics are subject to this interference. The quantitative coupling relationship is largely influenced by the type and dosage of anesthesia, including the actions on neural processing, vasoactive signal transmission, and vascular reactivity. The effects of anesthesia on the spatial gap between the neural and vascular response regions are not fully understood and require further attention to elucidate the mechanism of vascular control of CBF supply to the underlying focal and surrounding neural activity. The in-depth understanding of the anesthesia actions on neurovascular elements allows for better decision-making regarding the anesthetics used in specific models for neurovascular experiments and may also help elucidate the signal source issues in hemodynamic-based neuroimaging techniques

Vascular dimension, blood plasma speed and arteriovenous transit time in anesthetized rat cortex

The vascular dimensions and blood flow structure were systematically characterized along the branching orders of the cortical vessel trees in anesthetized rat cortex in vivo. The animals (300-340 g Sprague-Dawley rats, N = 4) were anesthetized with either isoflurane (~1.4%) or alpha-chloralose (45 mg/kg/h, i.v.). The portion of the skull over the somatosensory area was removed while the dura was kept intact. FITC-dextran (MW = 7000) was intravenously injected, and the cortical vasculature was visualized in vivo with confocal laser-scanning microscopy (488-nm excitation) or multi-photon excitation fluorescent microscopy (900-nm excitation). The lumen diameter of the vessels was measured from the arterial to venous compartments, including the capillaries distributed beneath the cortical surface at depths of up to~0.4 mm. The size of the major cerebral artery (A1) was 97 +/- 17 mum under isoflurane anesthesia, but was observed to decrease remarkably after injection of alpha-chloralose to 75 +/- 15 mum. Similar shrinkage of the lumen diameter was observed for the second (A2) and third (A3) branched arteries;66 and 45 mum under isoflurane vs. 44 and 35 mum under alpha-chloralose. However, no clear difference in lumen size for either anesthetic was found for the fourth (A4) and fifth (A5) branched arterioles (31 and 23 mum under isoflurane vs. 30 and 21 mum under alpha-chloralose), and all venous compartment vessels (V5, V4, V3, and V2); 22, 34, 85, and 182 mum under isoflurane, and 26, 35, 81, and 175 mum under alpha-chloralose, respectively. The results indicate that the relatively large size of the arteries and arterioles plays a major role in controlling baseline CBF during different anesthetic conditions. Further, with intravenously injected fluorescent microspheres (1 mum in diameter), the blood plasma speed and arteriovenous transit time were evaluated in each cerebrovascular compartment and across the various branching points. We observed similar arteriovenous transit times under either anesthetic condition and similar blood speed profiles. These findings further support the conclusion that the arterial regulation of blood volume(CBV) plays a key role in determining anesthetic-dependent baseline CBF.Neuroscience 2007, the 37th annual meetin