Blood–brain barrier preservation in the in vitro isolated guinea pig brain preparation

Abstract

The morphofunctional preservation of the blood-brain barrier (BBB) was evaluated in the isolated guinea pig brain maintained in vitro by arterial perfusion. Electron microscopy evaluation after 5 hr in vitro demonstrated that cerebral capillaries and BBB specializations in this preparation retain features compatible with structural integrity. BBB-impermeable and -permeable atropine derivatives arterially perfused to antagonize carbachol-induced fast oscillatory activity confirmed the functional preservation of the BBB in vitro. To study BBB function further, changes in extracellular K+ concentration during arterial perfusion of a high-K+ solution were measured with K+-sensitive electrodes positioned in the cortex and, as control, at the brain venous outlet, where the solution perfused through the brain arterial system was collected. After 5 hr in vitro, the [K+](o) values measured during high-K+ perfusion in the piriform and entorhinal cortices were 5.02 +/- 0.17 mM (mean +/- SE) and 5.2 +/- 0.21 mM, respectively (n = 6). Coperfusion of the high-K+ solution with the Na+/K+ pump blocker ouabain (10 microM; n = 4) induced consistently spreading depression preceded by a rise in [K+](o). Finally, sporadic, isolated spots of extravasation of the fluorescent marker fluorescein isothiocyanate (FITC)-dextran preferentially circumscribed to deep cortical layers was observed in brains perfused with FITC-dextran after 5 hr in vitro. The study demonstrates that the in vitro isolated guinea pig brain is viable for studying cerebrovascular interactions and BBB permeability of compounds active in the central nervous system.

Distribution of the Olfactory Fiber Input Into the Olfactory Tubercle of the In Vitro Isolated Guinea Pig Brain

“…*t-test, P Ͻ 0.05. preparation retains normal physiological properties close to the in vivo condition for several hours when perfused in vitro with a saline solution supplemented with a plasma expander (de Curtis et al 1991(de Curtis et al , 1994Biella et al 2002;Muhlethaler et al 1993). Moreover, the structural and functional preservation of the complex relationship between the vascular and the glial/ extracellular compartments that form the blood-brain barrier (BBB) has been shown in this preparation (Librizzi et al 2001;Mazzetti et al 2004). In this study, we used a combined electrophysiological and imaging approach to analyze the OT synaptic networks elicited by LOT electrical stimulation in guinea pig isolated whole brain.…”

Mechanisms of C-Reactive Protein-Induced Blood–Brain Barrier Disruption

“…To reproduce our findings in a system closer to the in vivo situation, we further investigated the effect of CRP in the in vitro isolated guinea pig brain preparation, in which the complex interactions between the vascular and the neuronal compartments are preserved. 17,27,28 Using this model, functional alterations in BBB can be monitored by measuring changes in the extracellular potassium concentration…”

Acute induction of epileptiform discharges by pilocarpine in the in vitro isolated guinea-pig brain requires enhancement of blood–brain barrier permeability

“…In order to evaluate the brain level of extracellular potassium ([K + ] o ), two-barrel glass pipettes were utilized to simultaneously record ion-selective signals and field responses (tip diameter, 3-5 μm) in EC and hippocampus (Librizzi et al, 2001). The conventional electrode barrel was filled with KCl 0.2 M. The pipette barrel utilized for [K + ] o measurements was filled at the tip with K + ionophore I cocktail A (Fluka, Buchs, Switzerland) after 1 min exposure to dimethyldichlorosylane vapors (Fluka, Steinheim, Germany) and was backfilled with 0.2 M KCl following a 2-h incubation at 120 °C.…”

“…We monitored the evoked response to paired LOT stimulation (interstimulus interval 40 ms) during histamine perfusion (n=5) and found no significant change in the ratio between the peak amplitude of mono-and disynaptic components of conditioned and conditioning responses in the PC (t-test, P<0.05). These data suggest that histamine is not modifying brain excitability in most of the experiments.We demonstrated that histamine was able to increase BBB permeability by analyzing the effects induced by simultaneous 10-min perfusion of pilocarpine (800 μM), histamine (100 μM) and atropine methyl bromide (5 μM), the BBB-impermeable derivative of atropine (Biesold et al, 1989;Librizzi et al, 2001). In all experiments (n=3) the presence of atropine methyl bromide prevented the development of pilocarpine-induced epileptiform activity and either reduced or abolished the oscillatory activity induced by a previous perfusion of pilocarpine (800 μM, 5 min; Fig.…”

“…The demonstration that histamine was, indeed, able to open BBB is provided by the experiments with atropine bromide, a BBB-impermeable compound (Biesold et al, 1989). If the BBB is functionally intact, atropine bromide does not block muscarinic-dependent activity (Librizzi et al, 2001). When BBB-opener histamine was co-perfused in the isolated brain, atropine bromide either hindered epileptiform discharges or substantially reduced fast oscillatory activity induced by pilocarpine.…”