Galanin antagonizes vasopressin-stimulated flank marking in male golden hamsters.

Microinjection of vasopressin (VP) into the anterior hypothalamus (AH) of golden hamsters induces a rapid bout of flank marking, a stereotyped scent marking behavior used for olfactory communication. In rats, VP is colocalized with galanin (GAL) in several brain regions. GAL has been shown to antagonize the postsynaptic actions of other cosecreted neurotransmitters including acetylcholine and norepinephrine; however, the ability of GAL to modulate the postsynaptic actions of VP has not been assessed. Here, we report that coadministration of GAL can block VP-induced flank marking in golden hamsters in a dose dependent manner. These findings provide the first evidence in any species that GAL can antagonize the central actions of VP. Using slice binding and receptor autoradiography, we have identified GAL binding sites in the AH and two other regions implicated in flank marking behavior (the lateral septum and central grey). These findings raise the possibility that endogenous GAL may function as an inhibitory modulator of this stereotypic scent marking behavior.

Leptin binding in the arcuate nucleus is increased during fasting.

The arcuate nucleus (ARC) mediates the anorexic effects of leptin and expresses the long form (Ob-Rb) of the leptin receptor. To determine whether ARC leptin binding increases when plasma leptin levels are low during fasting, [125I]-leptin specific binding to rat brain slices was measured by quantitative autoradiography. [125I]-leptin specific binding was dense in the ARC and increased 2-fold after a 48-h fast (P<0.001). These findings suggest that leptin receptor binding in the ARC is upregulated during fasting and that fasting changes the sensitivity of the ARC to leptin.

Vasopressin acts in the area postrema to attenuate the exercise pressor reflex in anesthetized cats.

Circulating arginine vasopressin (AVP) can enhance baroreflex function via its action in the area postrema (AP). We tested the hypothesis that AVP acts in the AP to enhance baroreflex function during static contraction and, in turn, attenuates the exercise pressor reflex. Thus mean arterial blood pressure (n = 9) and heart rate (HR) (n = 9) during 30 s of electrically stimulated hindlimb contraction were compared before and after bilateral microinjections of 200 nl of the AVP V1-receptor antagonist d(CH2)5Tyr(Me)-AVP (V1x) (1 ng/nl) into the AP of the anesthetized cat. This protocol was repeated in three other cats in which sinoaortic denervation (SAD) was performed before any intervention. Injection of V1x into the AP had no effect on baseline blood pressure or HR. However, pressor and HR responses to static contraction were augmented by 44 +/- 10 and 29 +/- 9%, respectively. Static contraction also increased plasma AVP from 15.9 +/- 2.0 to 25.5 +/- 3.4 pg/ml. In the SAD cats, microinjection of V1x had no effect on contraction-induced increases in blood pressure or HR. These results suggest that baroreflex opposition of the reflex cardiovascular response to static contraction is enhanced by the action of AVP in the AP.

Area postrema-induced inhibition of the exercise pressor reflex.

The exercise pressor reflex is opposed by the arterial baroreflex, and circulating peptides may act in the area postrema to enhance this inhibition. Therefore, we tested the hypothesis that the area postrema exerts an inhibitory effect on this reflex. Consequently, in six alpha-chloralose-anesthetized cats, blood pressure and heart rate responses to 30 s of electrically stimulated hindlimb contraction were compared before and after thermal coagulation of the area postrema. In six other cats, the same contraction-induced cardiovascular responses were assessed before and after chemical lesion of the area postrema using kainic acid (214 +/- 9 nl, 2.5-5 mM). Thermal lesion of the area postrema augmented blood pressure and heart rate responses to contraction from 29 +/- 5 to 47 +/- 7 mmHg (P < 0.05) and from 8 +/- 2 to 14 +/- 2 beats/min (P < 0.05), respectively. Chemical lesion of the area postrema enhanced contraction-evoked blood pressure (30 +/- 7 vs. 47 +/- 6 mmHg, P < 0.05) and heart rate (12 +/- 4 vs. 17 +/- 4 beats/min, P < 0.05) responses. These data suggest that the area postrema attenuates the exercise pressor reflex, possibly through the actions of circulating peptides on baroreflex function.

Production of hydroxyl radicals in contracting skeletal muscle of cats.

Reactive oxygen species increase during exhaustive contraction of skeletal muscle, but characterization of the specific species involved and their rates of production during nonexhaustive muscle contraction have not been investigated. We hypothesized that the production rate of hydroxyl radical (.OH) increases in contracting muscle and that this rate is attenuated by pretreatment with deferoxamine (Def) or dimethylthiourea (DMTU). We measured the rate of production of .OH before, during, and after 5 min of intermittent static contraction of the triceps surae muscles in cats (n = 6) using the formation of p-, m-, and o-tyrosines by hydroxylation of phenylalanine. L-Phenylalanine (30 mg/kg i.v.) was administered to each animal 3 min before contraction. Blood samples were collected from the popliteal vein 1 min before contraction; 1, 3, and 4.5 min during contraction; and 1 min after contraction. During and after contraction, the cumulative production rates of p-, m-, and o-tyrosines were elevated by 42.84 +/- 5.41, 0.25 +/- 0.04, and 0.21 +/- 0.03 nmol.min-1.g-1, respectively, compared with noncontracting triceps surae muscles. Pretreatment with Def (10 mg/kg i.v.; n = 5) or DMTU (10 mg/kg i.v.; n = 4) decreased the cumulative rates of production of p-, m-, and o-tyrosines during and after contraction. Additionally, the rate of tyrosine production increased in proportion to the percentage of maximal tension developed by the triceps surae muscles. These results directly demonstrate that .OH is produced in vivo in the skeletal muscle of cats during intermittent static contraction and that production can occur before the onset of fatigue.

Reactive oxygen species modify reflex cardiovascular responses to static contraction.

Reactive oxygen species can reflexly activate the cardiovascular system through stimulation of abdominal visceral afferents. The mechanism appears to involve hydroxyl radicals. We tested the hypothesis that reactive oxygen species contribute to the reflex cardiovascular response to static muscle contraction (i.e., the exercise pressor reflex). Thus blood pressure and heart rate responses to 5 min of intermittent electrically stimulated static contraction of the triceps surae muscles (15 s on, 15 s off) in anesthetized cats were compared before and after intravenous administration of the free radical scavengers dimethylthiourea (DMTU; 10 mg/kg; n = 8) or deferoxamine (Def; 10 mg/kg; n = 15). The contraction-induced pressor response was augmented from 51 +/- 6 to 61 +/- 7 mmHg after treatment with DMTU (P < 0.05) and from 44 +/- 8 to 58 +/- 8 mmHg after administration of Def (P < 0.05). Corresponding heart rate responses were not affected by either drug. Because this DMTU- or Def-induced augmentation of the exercise pressor reflex may have been due to a reduction in free radical-evoked vasodilation in the contracting skeletal muscle, popliteal artery blood velocity was measured with a Doppler flow transducer before and during contraction in the absence and presence of Def (n = 8). Blood velocity during contraction was not altered by Def (16 +/- 5 vs. 24 +/- 6 cm/s). These data suggest that reactive oxygen species exert an inhibitory effect on the exercise pressor reflex that is not associated with their local vasodilator properties. This response is opposite to that observed during stimulation of visceral afferents by reactive oxygen species.

Endogenous bradykinin in the thoracic spinal cord contributes to the exercise pressor reflex.

This investigation tested the hypothesis that bradykinin causes excitatory effects in the thoracic spinal cord that augment the exercise pressor reflex. Thus we performed 30 s of electrically stimulated static contraction of the hindlimb in the anesthetized cat (alpha-chloralose) to provoke reflex-induced increases in mean arterial pressure, maximal rate of rise of left ventricular pressure (dP/dt), and heart rate (i.e., the exercise pressor reflex). These three responses were compared before and 15 min after intrathecal injection of 2 micrograms (n = 3), 10 micrograms (n = 6), or 50 micrograms (n = 3) of the selective bradykinin B2- receptor antagonist HOE-140 into the thoracic spinal cord or 10 micrograms of this antagonist into the lumbar (n = 3) spinal cord. In three of the six cats in which 10 micrograms of HOE-140 were injected into the thoracic spinal cord, an additional contraction was performed 60-90 min after treatment. The 2-microgram dose of HOE-140 had no effect on the exercise pressor reflex. Injection of 10 micrograms of this antagonist into the thoracic spinal cord reduced the contraction-evoked pressor, maximal dP/dt, and heart rate responses by 49 +/-7, 58 +/- 4, and 64 +/- 13%, respectively (P < 0.05). Fifty micrograms of HOE-140 failed to attenuate these responses further. In the three cats in which an additional contraction was performed 60-90 min after treatment with 10 micrograms of the antagonist, blood pressure and dP/dt responses had returned, in part, toward initial values. Neither intravenous (n = 3) nor intrathecal injection of 10 micrograms of HOE-140 into the lumbar spinal cord had any effect on the contraction-induced cardiovascular responses. Thoracic injection of 50-200 ng of bradykinin provoked a pressor response of 26 +/- 5 mmHg that was abolished by a similar injection of 10 micrograms of HOE-140. These data suggest that endogenous bradykinin contributes to the exercise pressor reflex by an excitatory action in the thoracic spinal cord.

Spinal angiotensin II influences reflex cardiovascular responses to muscle contraction.

We tested the hypothesis that inhibition of angiotensin II (ANG II) AT1 receptors in the thoracic spinal cord attenuates the reflex cardiovascular response to electrically induced hindlimb static contraction (exercise pressor reflex). Consequently, in alpha-chloralose-anesthetized cats, contraction-induced increases in mean arterial blood pressure, maximal rate of rise in left ventricular pressure (dP/dt), and heart rate were compared before and after intrathecal injection of the AT1 receptor antagonist losartan (100 or 1,000 micrograms; n = 7). Losartan significantly diminished increases in blood pressure and maximal dP/dt provoked by static contraction by 33 +/- 5 and 31 +/- 6%, respectively. Conversely, these contraction-induced responses were unaffected by similar injection of ANG II into the lumbosacral spinal cord (n = 5). Moreover, intravenous injection of 100 micrograms losartan did not affect the cardiovascular response to contraction. Our data suggest that ANG II has a excitatory effect on the efferent arm of the exercise pressor reflex, which may be due to a facilitatory action on sympathetic nerve activity.