A physiological role for nitric oxide in the centrally mediated sympathetic and somatomotor ejaculatory response in anesthetized male Wistar rats.

Neurones in the lumbosacral spinal cord are known to play a significant role in ejaculation. In these same areas neurones containing nitric oxide synthase (NOS) have been described. This raised the question as to whether there is a physiological role for nitrous oxide (NO) in the spinal cord in sexual behavior. We first established immunohistochemical localization of NOS positive neurones in the lumbosacral spinal cord. NOS positive neurones were found in several areas of the lumbosacral cord. Namely the intermediolateral column (IML) at L(1)-L(4) and sacral cord; the dorsal gray commissure above the central canal at L(1)-L(2); the ventral gray area of lamina X below the central canal at L(3)-L(4); the superficial laminae of the dorsal horn at all levels. Secondly, we examined the role of NO in the generation of synchronized bursting activity within the vas deferens nerve and associated penile muscles, typical of sexual responses in the male anesthetized rat. NO modulators were applied directly to the spinal cord at T(13)-L(4) via a catheter placed subdurally (intrathecal) and their effect on the genital responses evoked by systemic administration of p-chloroamphetamine (PCA) or apomorphine (apo) (both 1 mg/kg) was observed. All responses evoked by PCA (n=4) or apo (n=3) were abolished or reduced (n=1) during intrathecal NOS inhibition using N((G)) nitro-L-Arginine methyl ester (l-NAME, 200 mM, 20-microl). NOS inhibition using l-NAME was reversed with simultaneous intrathecal application of the NO substrate l-arginine (100 mM, 20-microl, n=3). The selective neuronal NOS inhibitor 1-(2-trifluoro-methylphenyl) imidazole (100 mM, 20-microl, TRIM) also abolished all responses evoked by PCA (n=3). There was evidence that the responses within the vas deferens nerve (VDN) after PCA or apo were enhanced following NO donation using sodium nitroprusside (SNP, 1 mM, 20-microl). Furthermore, a PCA-like response within the VDN was evoked following intrathecally applied l-glutamic acid (200 mM, 20-microl) in six of 26 animals and also by intrathecal SNP in two of eight animals. In conclusion the results suggest a significant excitatory role for NO in the bursting pattern of synchronized discharge generated in autonomic and somatic outflows from the lumbosacral cord by neurones governing ejaculation.

Neuronal excitability in the periaqueductal grey matter during the estrous cycle in female Wistar rats.

Extracellular recordings were made from output neurons in the dorsal half of the periaqueductal gray matter (dPAG) in urethane-anesthetized female Wistar rats. All the neurons were quiescent. A basal level of firing was therefore induced by continuous iontophoretic application of D,L-homocysteic acid (DLH). In the presence of the GABA(A) receptor antagonist bicuculline methiodide (BIC 0-30 nA) the DLH-induced firing increased further, revealing the presence of ongoing GABAergic inhibitory tone on the recorded neurons. The BIC-induced increase in firing rate was significantly greater in neurons recorded during estrus (Est) and late diestrus (LD) compared with proestrus (Pro) and early diestrus (ED) suggesting that GABAergic tone was lower in Est and LD. I.v. injection of the panicogenic cholecystokinin (CCK)(B) receptor agonist pentagastrin (PG, 40 microg kg(-1)) produced an increase in firing rate in 12/17 (70%) of neurons tested in the dPAG. Iontophoretic application of PG (10-30 nA) also produced a current-related increase in firing rate in 73.6% of the neurons tested. The excitatory response was reduced during application of the selective CCK(B) receptor antagonist beta-[2-([2-(8-azaspiro[4.5]dec-8-ylcarbonyl)-4,6-dimethylphenyl]amino)-2-oxoethyl]-(R)-napthalenepropanoic acid (CR2945) (60 nA, n=6). The PG-evoked increase in firing rate was significantly greater in neurons recorded during Est and LD compared with during Pro and ED. Juxtacellular labeling with neurobiotin in eight neurons revealed multipolar cells 12-44 microm diameter with up to six primary dendrites. In three of eight neurons, a filled axon was present and coursed without branching toward the perimeter of the periaqueductal gray matter (PAG). The estrous cycle-related change in responsiveness to BIC and PG suggests that the panic circuitry in the PAG may become more responsive to panicogenic agents during estrus and late diestrus as a consequence of a decrease in the intrinsic level of inhibitory GABAergic tone. The findings may have implications for understanding the neural processes that underlie the development of premenstrual dysphorias in women.

Cardiovascular and respiratory responses to a panicogenic agent in anaesthetised female Wistar rats at different stages of the oestrous cycle.

In urethane-anaesthetised female Wistar rats, intravenous injection of the panicogenic CCK(B) receptor agonist pentagastrin (0.002-80 microg/kg) evoked a dose-related increase in blood pressure, heart rate and ventilation. The response was blocked in the presence of the selective CCK(B) receptor antagonist CR2945 (1 mg/kg i.v.). The same pattern of cardiovascular and respiratory changes was evoked by microinjection of pentagastrin (0.3 nmol in 250 nL) into the dorsal half of the periaqueductal grey matter (PAG). The effect of intra-PAG administration of pentagastrin was also abolished following injection of CR2945 (1 mg/kg, i.v.). Responsiveness to systemically administered pentagastrin was enhanced in rats in late dioestrus. At the highest dose tested (80 microg/kg), the pressor response, tachycardia and tachypnoea evoked in rats in late dioestrus was significantly higher than rats in proestrus. For rats in oestrus, the pressor response and tachycardia but not tachypnoea were also significantly larger than the response evoked in rats in early dioestrus. The results suggest that the dorsal half of the PAG (dPAG) plays a key role in mediating the cardiovascular and respiratory responses evoked by systemically administered CCK(B) agonists. The enhanced responsiveness to panicogenic agents during late dioestrus may be related to changes in the functional responsiveness of gamma-aminobutyric acid (GABA)ergic circuitry in the dPAG due to plasticity of GABA(A) receptor subunit expression as a consequence of falling progesterone levels.

Effects of direct sympathetic and vagus nerve stimulation on the physiology of the whole heart--a novel model of isolated Langendorff perfused rabbit heart with intact dual autonomic innervation.

A novel isolated Langendorff perfused rabbit heart preparation with intact dual autonomic innervation is described. This preparation allows the study of the effects of direct sympathetic and vagus nerve stimulation on the physiology of the whole heart. These hearts (n = 10) had baseline heart rates of 146 +/- 2 beats x min(-1) which could be increased to 240 +/- 11 beats x min(-1) by sympathetic stimulation (15 Hz) and decreased to 74 +/- 11 beats x min(-1) by stimulation of the vagus nerve (right vagus, 7 Hz). This model has the advantage of isolated preparations, with the absence of influence from circulating hormones and haemodynamic reflexes, and also that of in vivo preparations where direct nerve stimulation is possible without the need to use pharmacological agents. Data are presented characterising the preparation with respect to the effects of autonomic nerve stimulation on intrinsic heart rate and atrioventricular conduction at different stimulation frequencies. We show that stimulation of the right and left vagus nerve have differential effects on heart rate and atrioventricular conduction.