Alterations In Membrane Potential Of Arterial Smooth Muscle Cells And Ca_L Currents Of Cerebrovascular Smooth Muscle Cells In 2-week Hind-limb Unweighting Rats

The occurrence of post spaceflight cardiovascular dysfunction (PFCD) is related to multiple mechanisms which are not fully understood. Previous works found that: weightlessness simulation can induce enhanced myogenic tone and increased vasoreactivity, hypertrophy, and perivascular hyperinnervation in rat cerebral arteries, whereas can induce attenuated myogenic tone and vasoreactivity, atrophy, and perivascular hypoinnervation in rat hindquarter arteries. Based on recent results, the"peripheral effector mechanism hypothesis"has been raised to explain the inability to elevate the peripheral vascular resistance adequately and altered autoregulation of cerebral vasculature during orthostatic challenges in astronauts and cosmonauts postflight. Our recent work further showed that the cellular and molecular mechanisms regarding vascular adaptation to microgravity may involve vascular local renin-angiotensin system (L-RAS) and the ion channel remodeling mechanism of vascular smooth muscle cells (VSMCs). Study on countermeasures of simulated weightlessness suggests that enhanced myogenic tone and increased vasoreactivity are two independent procedures in cerebral vascular wall during microgravity exposure. To sum up, we think the VSMCs ion channel remodeling mechanism maybe involved and play a role in differential arteries adaptation changes in rats subjected to microgravity environment. Our previous work have reported that the L-type voltage-dependent calcium channel (CaL) current densities of VSMCs isolated from rats cerebral arteries have tendency of increasing after a 3-day weightlessness simulation, whereas the current densities and expression of CaL protein increased significantly after a 4-week weightlessness simulation. But further studies are required to determine the time course feature of CaL function changes during 4-week weightlessness simulation.Membrane potential (MP) and intracellular calcium concentration ([Ca2+]i) are two important factors in regulating VSMCs contraction and relaxation. A small change of MP could result in significant alteration in vascular tension. MP is normally not a result of single ion channel changes, but a comprehensive change of several ion channels. So choosing MP to investigate the vasoreactivity of VSMCs is more objective than choosing single ion channel. Study on spontaneous hypertensive rat (SHR) revealed that the MP of basilar artery VSMCs is higher than the result of Wistar rat, and vasoreactivity increased. As previous discussed, the cerebral arteries of rat subjected to simulated weightlessness have a similar adaptation change as that of SHR. Study on renal hypertensive rat suggests that membrane depolarization is a potential signal that may contribute to the development of abnormal vascular tone. But influences of weightless simulation on MP of VSMCs are still unrevealed, and so dose the MP mechanism in development of vasoreactivity changes in rat subjected to microgravity.To investigate potential role of MP changes and ion channel remodeling mechanism in vasoreactivity changes, we did experiments as below:1. We used intracellular microelectrode technique and tail-suspended rat model to investigate the MP of VSMCs isolated from several arteries of normal rats, and compared with the results from rats subjected to a 2-week simulated weightlessness, especially focused on the MP changes of cerebral arteries and vasoreactivity changes to drugs such as KCl, PE and Ach.2. We used whole-cell recording patch clamp technique and tail-suspended rat model to investigate alterations of CaL in cerebral vascular smooth muscle cells isolated from rats subjected to a 2-week simulated weightlessness, and influence of Bay K 8644 (an agonist of CaL) to the channel currents.The main findings of the present work are as follows:1. The MP of basilar artery VSMCs isolated from 2-week simulated weightlessness rat is -49.54±6.80 mV (n=35), which is about 12 % higher than the result of control group rat (-56.03±8.45 mV, n=29; P<0.01). The MP of common carotid artery VSMCs isolated from 2-week simulated weightlessness rat is -51.50±6.09 mV (n=23), which is about 12 % higher than the result of control group rat (-59.12±6.69 mV, n=19; P<0.05). The vasoreactivity of basilar artery and common carotid artery to drugs such as KCl and PE increases in a 2-week simulated weightlessness rats compared with result of control group rats. Ach have no significant changes on VSMC MP in every artery in our experiment.2. Cerebral VSMCs CaL current density increased significantly in 2-week simulated weightlessness rats compared with that of simultaneous control group rats, current density was (-7.2±0.4 pA/pF) vs (-5.0±0.3 pA/pF)(P<0.01, n=28) respectively. After Bay K 8644 added, compared with that of simultaneous control group rats, the amplification of CaL current density in 2-week simulated weightlessness rats was also higher, which was (-10.7±1.3 pA/pF) vs (-4.7±0.5 pA/pF)(P<0.01, SUS, n=17; CON, n=16) respectively. But membrane capacitance (Cm), access resistance (Ra), and other parameters of CaL such as steady state activation / inactivation curves have no significant changes compared with those of control group.To sum up, 2-week simulated weightlessness increase VSMCs CaL current density significantly in cerebral arteries of rats, which testify the change of CaL function last in the whole weightless simulation course, and the MP change after weightless simulation suggest MP mechanism was involved in development of adaptation changes in cerebral arteries induced by weightless simulation.