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. 1998 Nov 1;512(Pt 3):927–938. doi: 10.1111/j.1469-7793.1998.927bd.x

Enhanced vasoconstrictor responses in renal and femoral arteries of the golden hamster during hibernation

Parastoo Karoon 1, Gillian Knight 1, Geoffrey Burnstock 1
PMCID: PMC2231249  PMID: 9769433

Abstract

  1. The present study assessed local regulation of vascular tone of euthermic (control), cold control and hibernating golden hamsters. Sympathetic neurotransmission in the renal artery, the long term effects of hibernation on perivascular nerve activity, and the responsiveness of femoral artery to a number of neurotransmitters and hormones with both constrictor and dilator actions during hibernation are described.

  2. The contractile responses of the renal arterial rings to transmural nerve stimulation (80 V, 0.1 ms, 4–64 Hz, for 1 s) were negligible in controls, significantly increased at higher frequencies of stimulation in cold controls and markedly enhanced in the hibernating group at all frequencies tested. The contractile responses to exogenous noradrenaline (NA; 0.1–100 μm) were significantly increased in the renal arteries of hibernating hamsters compared with controls, but not compared with cold controls. Responses to exogenous ATP (1–3000 μm) and KCl (120 mm) were similar among all experimental groups.

  3. The maximal contractile responses of femoral arterial rings to the sympathetic co-transmitter ATP and 5-hydroxytryptamine were increased by approximately 124 % and 99 %, respectively, in hibernating compared with cold control preparations without a change in the concentration of agonist that produces half-maximal response. However, the responses to NA were not altered during hibernation.

  4. Vasoconstriction of femoral arterial rings in response to arginine vasopressin was significantly enhanced in both cold controls and hibernating groups, while vasoconstriction in response to endothelin-1 and KCl was unaltered.

  5. The dilator responses of femoral arterial rings to acetylcholine, sodium nitroprusside and adenosine were not different among the groups.

  6. It is suggested that the marked augmentation of sympathetic neurotransmission, selective supersensitivity of the vascular smooth muscle to sympathetic contractile agents and unaltered vasodilatory mechanisms may provide a means for maintenance of vascular tone and peripheral resistance during hibernation.

Onset of hibernation is accompanied by a decline in heart rate and consequent decrease in systolic and diastolic blood pressure. Despite this fall, the mean arterial pressure remains within the range observed in normal conscious animals as a result of an increase in peripheral resistance (Lyman & O'Brien, 1960; Wells, 1971; Zatzman, 1984). Lyman & O'Brien (1963) suggested that although some of the changes observed in the blood flow were attributable to a temperature-dependent increase in blood viscosity an increase in vascular tone was also a contributing factor. Maclean (1981) reported that several characteristics of blood of hibernators combined to make it unlikely that an increase in blood viscosity was a major contributor to the observed increase in the peripheral resistance. The latter argument was further supported by work of Miller et al. (1986) who showed that responsiveness to noradrenaline (NA) was increased in renal vessels of hibernating compared with non-hibernating woodchucks, Marmota monax. This adaptive response was specific to hibernating animals and led to the hypothesis that changes in vascular responsiveness contribute to the regional control of blood flow in hibernating animals.

During hibernation renal function is reduced to 10 % of its euthermic level in many hibernating species. The marmot and the bear demonstrate an increase in renal vascular resistance indicated by increased filtration fraction (Brown et al. 1971; Zatzman & South, 1972).

The vasculature is under dual local control by perivascular nerves and endothelial cells, which involves a plethora of regulatory substances including monoamines, purines, peptides and nitric oxide (NO) (Ralevic & Burnstock, 1995). The extensive influence of sympathetic perivascular nerves on the regional control of blood flow in many mammals is well established. Evidence has accumulated that NA, ATP and neuropeptide-Y (NPY) act as co-transmitters in perivascular sympathetic nerves (Burnstock, 1995). 5-Hydroxytryptamine (5-HT) is taken up and released from perivascular sympathetic nerves, and activates its postjunctional receptors. However, 5-HT is not synthesized in these nerves and is termed a ‘false transmitter’ (Jackowski et al. 1988). The control exerted by each of these substances differs considerably in different blood vessels and species (Burnstock, 1995). Sympathetic neurotransmission has been described in the renal vasculature of rat, rabbit and dog (Petrovic et al. 1988; Luff et al. 1991). Although the exact nature of sympathetic neurotransmission in the femoral artery has not been described before, adrenergic nerve fibres and related terminals were found in the femoral artery of dog (Amenta et al. 1979). Additionally, NA has been localized within the wall of the femoral artery of rat and pig (Dolezel et al. 1975). The involvement of the sympathetic nervous system and its co-transmitters on the regional control of renal and femoral blood flow in hamster has not been described, nor its role in the local control of vascular tone during hibernation and/or cold exposure.

The influence of endothelial cells in local control of vasculature has also been well documented. Release of vasodilators, including NO, and vasoconstrictors, including endothelins, from endothelial cells has been reported (Palmer et al. 1987; Emori et al. 1989). There is also evidence for storage of arginine vasopressin (AVP) in endothelial cells (Loesch et al. 1991). The magnitude of control exerted by each of these substances differs considerably in different blood vessels (see Ralevic & Burnstock, 1995). So far, there has been no study to address endothelium-mediated responses during hibernation or cold exposure. Nor has there been any investigation on the effect of hibernation and cold exposure on responses to endothelin-1 (ET-1) and AVP.

The present study investigates factors influencing local control of vascular tone in age-matched controls (controls), cold-exposed controls and hibernating golden hamsters. To this end, isolated renal and femoral arterial rings were used to evaluate the sympathetic neurotransmission and test for adrenergic-purinergic perivascular co-transmission as well as the endothelium-dependent and -independent actions of vasodilator agents.

METHODS

Animal treatment

Syrian or golden hamsters (Mesocricetus aurartus; 125–150 g) were used; they are ‘permissive hibernators’ since they may or may not hibernate depending on hoarded food stores and external environmental conditions (Morrison, 1960). Hibernation in this species can be induced by manipulating photoperiod or temperature and creating pseudo-winter conditions in the laboratory at any time of the year (Hoffman et al. 1965). Male hamsters were caged in groups of five in a refrigerated incubator (Leec Ltd, Nottingham, UK) initially set at 20°C and with the light:dark period set at 8:16 h. The period of light was reduced by 30 min daily, together with a gradual temperature reduction to reach the final condition of 2 h light each day and an ambient temperature of 9°C. This acclimatization procedure took 7–10 days.

The animals were then transferred to individual cages in a cold room which was set at 5°C with 2 h of light per day with food and water ad libitum. Inspection of the animals outside the designated light period was only performed under a 10 W red photographic light. To check whether the animals were hibernating, sawdust was sprinkled on their backs (which would waken a sleeping but not a hibernating hamster). Animals were in hibernation for 8 weeks, with periodic arousal, but were killed only after a deep hibernation bout of 1–4 days. Animals which underwent similar treatment but did not hibernate were used as cold controls. Age- (3 months) and sex-matched animals kept at room temperature were used as controls. Two separate sets of hamsters, undergoing the above procedure, were used for the studies of the renal and femoral arteries.

In vitro pharmacology

The hamsters were killed in accordance with UK Home Office guidelines by a rising concentration of CO2. The cheek and rectal temperatures were measured immediately after asphyxiation. Renal and femoral arteries were dissected and cleaned of excess tissue. Ring segments of 3–4 mm in length were cut and mounted in 5 ml organ baths containing (mm): 133 NaCl, 4.7 KCl, 1.35 NaH2PO4, 16.3 NaHCO3, 0.61 MgSO4, 2.52 CaCl2 and 7.8 glucose, gassed with 95 % O2-5 % CO2 at 37°C. An initial resting tension of 1 g was applied to the arterial rings, which were then left to equilibrate for 60–90 min. At the end of this period the tension on the vessel was taken as the resting tension and no further mechanical adjustment was made during experimentation. Isometric tension was recorded with a Grass FT03C transducer and displayed on a Grass ink-writing polygraph (model 79).

In renal arterial rings, transmural nerve stimulation (TNS) was achieved by passing a current between two electrodes parallel to the arterial rings. Parameters for TNS (80 V, 0.1 ms, 4–64 Hz, for 1 s) were selected, since they evoke both adrenergic and purinergic components of perivascular sympathetic neurotransmission in similar vascular preparations (Karoon et al. 1995). Reproducible vascular responses were obtained with 1 s duration of TNS. Prazosin was added to the bath 20 min prior to construction of a frequency-response curve, then prazosin together with α,β-methylene ATP was added to the bath for a further 20 min and another frequency-response curve established. Desensitization to α,β-methylene ATP was confirmed by no response to a repeated dose of the agonist. Cumulative concentration-response curves for NA and ATP were established.

In femoral arterial rings cumulative concentration-response curves for NA, ATP, 5-HT, AVP and ET-1 were established at basal tone. Vasodilator responses were assessed in preparations where the tone was raised by a concentration of NA that would give 50 % of maximal response (EC50). When the contraction had plateaued, cumulative concentration-response curves were constructed for acetylcholine (ACh), adenosine and sodium nitroprusside (SNP). Relaxant responses are expressed as percentage relaxation of the NA-induced contraction.

Contractile responses in both renal and femoral arterial preparations were evaluated as an increase in tension (g). In all preparations contractile response to 120 mm KCl was measured.

Drugs used

Noradrenaline bitartrate, Na2ATP, prazosin, α,β-methylene ATP, tetrodotoxin, 5-hydroxytryptamine, acetylcholine bromide, adenosine and sodium nitroprusside were obtained from Sigma Chemical Company. Arginine vasopressin and endothelin-1 were purchased from Cambridge Research Biochemicals (Northwich, UK). Monoamines were dissolved in ascorbic acid (0.1 mm); all other drugs were dissolved in distilled water.

Data analysis

All data are expressed as means ± s.e.m. The pD2 values were calculated as the negative log of the concentration required to produce 50 % of maximal response. Responses to ET-1 did not reach a maximum, and so pD75 values (negative log of the concentration required to produce vasoconstriction of 0.75 g) were calculated. Statistical analysis was performed using analysis of variance (ANOVA) and Student's paired or unpaired t test as appropriate for individual points and one-way ANOVA for the entire curve, using Instat (GraphPad Software, Inc., San Diego, CA, USA). A value of P less than 0.05 was considered statistically significant.

RESULTS

Animal treatment

Hamsters started hibernating after 10 weeks in a cold room and were allowed to hibernate for a further 8 weeks. During this time, bouts of 3–5 days of deep hibernation occurred. The rates of hibernation were 42 % and 48 % for those animals from which renal and femoral arteries were taken, respectively. Little is known as to why some hamsters maintained under conditions conducive to hibernation fail to hibernate. However, preparation for hibernation is accompanied by altered endocrine function (Hoffman et al. 1965) which may differ among individual hamsters. Hibernation is also genetically controlled (Karmanova, 1995) and may have been inadvertently bred out of the population of laboratory hamsters that are commercially available.

There was a significant weight loss in both hibernating and cold controls compared with the control group (Table 1). Cheek pouch and rectal body temperatures were also significantly reduced in hibernating animals compared with both cold control and control groups (Table 1). Two separate sets of hamsters were used for the study of renal and femoral artery and the mean values of body weights, cheek pouch and rectal temperatures are given in Table 1A and B, respectively.

Table 1.

Body weight, and cheek pouch and rectal temperatures of hamsters after 8 weeks hibernation

A. Renal artery studies B. Femoral artery studies
Hibernating Cold control Control Hibernating Cold control Control
Weight (g) 107.9 ± 5.1 (5)* 138.6 ± 5.5 (7)* 162.7 ± 8.7 (5) 105.8 ± 1.0 (6)*** 130.5 ± 4.1 (7) *** 151.0 ± 2.9 (10)
Cheek pouch temp. (°C) 9.1 ± 0.3 (5)*** 35.1 ± 0.5 (7) 35.4 ± 0.4 (5) 14.2 ± 1.9 (6)*** 33.0 ± 0.3 (7) *** 36.1 ± 0.2 (10)
Rectal temp. (°C) 10.0 ± 0.5 (5)*** 31.9 ± 0.7 (7) 32.0 ± 0.6 (5) 12.2 ± 1.4 (6)*** 32.4 ± 0.7 (7)* 34.0 ± 0.1 (10)
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Data are means ±s.e.m. (number of animals).

*

P < 0.05 vs. control

***

P < 0.001 vs. control.

Pharmacology

In golden hamster renal arterial rings, TNS (4–64 Hz) evoked frequency-dependent contractile responses of different magnitude in the three experimental groups (Fig. 1). These responses were abolished by application of tetrodotoxin (1 μm), thus revealing their neural origin. In the euthermic state (control group), contractile responses to TNS were only detectable at higher frequencies of stimulation and during cold exposure (cold control group) these responses were significantly greater. However, in the hibernating group contractile responses to TNS were also evident at lower frequencies and were significantly higher than both control and cold control groups at all frequencies tested (Figs 1 and 2). At 64 Hz vascular responses were 0.27 ± 0.06 g (n = 5), 0.10 ± 0.02 g (n = 7) and 0.05 ± 0.01 g (n = 5) in hibernating, cold control and control animals, respectively (Fig. 2).

Figure 1. Frequency-response recordings for transmural nerve stimulation.

Figure 1

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Recording of a typical experiment showing vasoconstrictor responses to transmural nerve stimulation (80 V, 0.1 ms, 4–64 Hz, for 1 s) in hamster renal arteries taken from control (top panel), cold control (middle panel) and hibernating (bottom panel) animals.

Figure 2. Frequency-response relationship in hamster renal arteries.

Figure 2

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Frequency-response curves to transmural nerve stimulation (80 V, 0.1 ms, 4–64 Hz, for 1 s) taken from controls (•, n = 5), cold controls (○, n = 7) and hibernating hamsters (□, n = 5). *P < 0.05, **P < 0.01vs. control; †P < 0.05vs. cold control.

In the presence of 1 μm prazosin, vascular responses to TNS were reduced in all three groups tested (Fig. 3). Furthermore, exposure of preparations to the desensitizing agonist of P2X receptors, α,β-methylene ATP (1 μm), produced rapid transient contractions in all the preparations tested, after which the residual component of TNS was not significantly reduced (Fig. 3).

Figure 3. Different components of frequency-response relationship in hamster renal arteries.

Figure 3

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Frequency-response to transmural nerve stimulation (80 V, 0.1 ms, 4–64 Hz, for 1 s) in the absence (▪) and in the presence of prazosin (1 μm, Inline graphic), and prazosin (1 μm) plus α,β-methylene ATP (1 μm, Inline graphic) taken from controls (n = 5) (A), cold controls (n = 7) (B) and hibernating hamsters (n = 5) (C).

In hibernating animals the noradrenergic component of perivascular sympathetic neurotransmission was greater than in both control and cold control groups with statistical significance being reached at 8, 16, 32, 50 and 64 Hz. In cold control animals the noradrenergic component of perivascular sympathetic neurotransmission was greater than in control animals at 50 and 64 Hz.

Application of exogenous NA (0.1–100 μm) to the renal arterial rings evoked concentration-dependent contractile responses which were significantly greater in hibernating than in control animals (Fig. 4A). However, there was no significant difference between hibernating and cold control animals. NA-evoked maximal responses were significantly greater in hibernating animals than in control animals, but did not differ between hibernating and cold control animals (Table 2A). No significant difference was seen among the pD2 values of the three experimental groups (Table 3A).

Figure 4. Concentration-response relationship for exogenous neurotransmitters in hamster renal artery.

Figure 4

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Concentration-response curves showing vasoconstrictor responses to noradrenaline (NA, A) and ATP (B) of arterial rings taken from controls (•, n = 6), cold controls (○, n = 7) and hibernating animals (□, n = 4). *P < 0.05vs. control.

Table 2.

Maximum contractile response (g) or dilatory response (percentage relaxation) to vasoactive agents

A. Renal artery B. Femoral artery
Vasoactive agent Hibernating Cold control Control Hibernating Cold control Control
NA (g) 1.93 ± 0.21 (4) ** 1.66 ± 0.21 (7) 1.13 ± 0.23 (6) 1.95 ± 0.15 (4) 1.38 ± 0.17 (5) 1.62 ± 0.26 (5)
ATP (g) 0.85 ± 0.14 (4) 0.68 ± 0.15 (7) 0.59 ± 0.15 (6) 1.55 ± 0.11 (3)** 0.69 ± 0.11 (3) 0.81 ± 0.11 (4)
KCl (g) 1.65 ± 0.06 (5) 1.48 ± 0.14 (5) 1.33 ± 0.27 (5) 1.66 ± 0.21 (4) 1.27 ± 0.15 (6) 1.48 ± 0.16 (5)
5-HT (g) — — — 1.83 ± 0.16 (4)** 0.92 ± 0.15 (5) 0.99 ± 0.10 (4)
ET-1 (g)† — — — 1.62 ± 0.19 (4) 1.46 ± 0.18 (6) 1.32 ± 0.13 (3)
AVP (g) — — — 1.42 ± 0.26 (4) 1.12 ± 0.13 (5) 0.99 ± 0.07 (5)
ACh (% relaxation) — — — 81.0 ± 11.4 (4) 79.4 ± 13.9 (6) 80.2 ± 9.9 (4)
SNP (% relaxation) — — — 90.6 ± 8.5 (4) 100.0 ± 0.0 (6) 99.7 ± 0.3 (5)
Adenosine (% relaxation) — — — 74.6 ± 14.6 (4) 74.1 ± 9.7 (6) 64.5 ± 12.6 (4)
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†

Vascular response to the highest ET-1 concentration tested (0.3 mm). Data are means ± s.e.m. (number of observations).

**

P < 0.01vs. control.

Table 3.

pD2 and pD75 values

A. Renal artery B. Femoral artery
Vasoactive agent Hibernating Cold control Control Hibernating Cold control Control
NA (pD2) 5.63 ± 0.13 (4) 5.58 ± 0.10 (7) 5.71 ± 0.15 (6) 6.31 ± 0.16 (4) 6.09 ± 0.12 (5) 6.20 ± 0.14 (5)
ATP (pD2) 3.75 ± 0.13 (4) 3.51 ± 0.12 (7) 3.74 ± 0.23 (5) 3.69 ± 0.14 (4) 3.69 ± 0.15 (4) 3.96 ± 0.10 (5)
5-HT (pD2) — — — 6.78 ± 0.30 (4) 6.53 ± 0.17 (5) 5.88 ± 0.07 (3)
ET-1 (pD75) — — — 8.00 ± 0.23 (4) 7.83 ± 0.21 (6) 7.58 ± 0.22 (4)
AVP (pD2) — — — 10.53 ± 0.21 (4)** 10.48 ± 0.30 (5)* 9.51 ± 0.09 (5)
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pD2, negative log of concentration of agonist producing half-maximal response; pD75, negative log of concentration of agonist producing response of 0.75 g tension. Data are means ±s.e.m. (number of observations).

*

P < 0.05vs. control

**

P < 0.01vs. control.

Vasoconstrictor response of the renal artery to ATP (1–3000 μm) was not significantly altered in either the hibernating animals or cold controls compared with control animals (Fig. 4B). The maximal contractile response and the pD2 values did not differ significantly among the three experimental groups (Tables 2A and 3A).

In the femoral artery, vasoconstrictor responses to NA (0.01–100 μm) were not significantly altered (Fig. 5A). Maximal contractile response and the pD2 values did not differ significantly among the three experimental groups (Tables 2B and 3B). In contrast, vasoconstriction in response to ATP (1 μm to 10 mm) was significantly enhanced in preparations from hibernating animals at concentrations of 1–10 mm (Fig. 5B). Maximal constriction in hibernating animals was approximately 125 % greater than that of the cold control preparations and approximately 91 % greater than that of the controls (Table 2B). There was no significant difference in pD2 values (Table 3B).

Figure 5. Concentration-response relationship for contractile neurotransmitters in hamster femoral artery.

Figure 5

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Concentration-response curves showing vasoconstrictor responses to NA (A), ATP (B) and 5-HT (C) in isolated arterial rings taken from hibernating (□), cold control (○) and control (•) hamsters. All results are expressed as means ± s.e.m. n = 4–5. *P < 0.05, **P < 0.01vs. control.

Vasoconstrictor responses to 5-HT in femoral artery preparations from hibernating animals were also significantly enhanced at concentrations of 0.3–30 μm (Fig. 5C). Maximal constriction in hibernating animals was approximately 99 % greater than that of the cold control preparations and approximately 85 % greater than that of the controls (Table 2B). There was no significant difference in pD2 values (Table 3B).

In the femoral artery, concentration-response curves of both hibernating and cold control groups for AVP (0.01- 300 nm) were significantly shifted to the left (Fig. 6A). The pD2 values were increased in the hibernating group and cold controls relative to the control group (Table 3B). There was no significant difference in maximal constriction among the groups (Table 2B).

Figure 6. Concentration-response relationship for contractile peptides in hamster femoral artery.

Figure 6

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Concentration-response curves showing vasoconstrictor responses to AVP (A) and ET-1 (B) in isolated arterial rings taken from hibernating (□), cold control (○) and control (•) hamsters. All results are expressed as means ± s.e.m. n = 4–6.

The concentration-response curve for ET-1 in the femoral artery did not reach a plateau in any of the experimental groups (Fig. 6B). Thus, pD75 values (negative log of concentration of agonist producing a response of 0.75 g tension), rather than pD2 values, were calculated. Vasoconstrictor response to the highest concentration of ET-1 tested (0.3 μm) and the pD75 values were unaltered among the groups (Tables 2B and 3B).

Contractile tension developed by 120 mm KCl in both renal and femoral arteries did not differ significantly among the three groups of animals tested (Table 2A and B).

ACh produced concentration-dependent sustained relaxations of hamster femoral arteries pre-contracted with an EC50 concentration of NA. Neither the concentration-response curves nor the maximal relaxations in response to ACh, SNP or adenosine differed among the groups (Fig. 7A, B and C, respectively; Table 2B).

Figure 7. Concentration-response relationship for vasodilator agents in hamster femoral artery.

Figure 7

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Concentration-response curves showing vasodilator responses to ACh (A), SNP (B) and adenosine (C) in isolated arterial rings precontracted by NA, taken from hibernating (□), cold control (○) and control (•) hamsters. All results are expressed as means ± s.e.m. n = 4–6.

DISCUSSION

The present results support the general hypotheses that (1) during hibernation and to a lesser extent cold acclimatization periarterial sympathetic neurotransmission is augmented, (2) hibernation induces selective supersensitivity of vascular smooth muscle to ATP and 5-HT that is independent of cold acclimatization, (3) long term exposure to cold results in an altered postjunctional action of NA in renal and supersensitivity to AVP in the femoral arterial preparations, and (4) hibernation and cold acclimatization have no influence on endothelium-dependent or -independent vasodilatory mechanisms.

The present study demonstrates for the first time that sympathetic neurotransmission in the renal artery of the golden hamster differs in euthermic, cold-exposed and hibernating conditions. Contractile responses to TNS were only elicited at high frequencies of stimulation in the euthermic state. Renal periarterial nerve stimulation induces vasoconstriction at frequencies as low as 2 Hz in rat and rabbit (Sehic et al. 1994). At present, with no other data available on the responsiveness of renal arterial preparations to TNS, we can offer no explanation for the difference in minimum frequencies that elicit responses between hibernating and non-hibernating species. In the present study, the response to TNS was significantly increased in the cold environment, whilst an augmentation of responses to nerve stimulation was seen at all the frequencies tested during hibernation.

Vasoconstriction of renal arterial rings evoked by TNS was largely antagonized by the α1-adrenoceptor antagonist prazosin, thus indicating that sympathetic neurotransmission in hamster renal artery is mediated mainly by NA through α1-adrenoceptors. Since a small residual response to TNS was left after blockade of α-adrenoceptors, the possibility of purinergic co-transmission was examined. In the present study, the residual response to TNS left after blockade of the adrenergic transmission was reduced by desensitization of P2X receptors with α,β-methylene ATP, but this was not statistically significant. Therefore, if there is a purinergic component of sympathetic transmission it is very small or a P2X receptor subtype distinct from the classical P2X1 subtype, insensitive to desensitization by α,β-methylene ATP, may be involved. Such a receptor has been recently reported to mediate ATP-evoked vasoconstriction of the rat renal and pulmonary vascular beds (Rubino & Burnstock, 1996).

Responses to exogenous NA in the renal arterial rings were enhanced in hibernating animals compared with the control group, although by only approximately one-third as much as those to TNS. This enhancement was likely to be a consequence of exposure to cold since responses in the hibernating group were not significantly greater than those of the cold control group. Other studies have provided diverse evidence on responsiveness of the vasculature to NA during long term cold exposure and hibernation. During cold exposure, reports indicate no changes in the responses of aorta, renal and femoral arterial preparations of winter woodchucks (Miller et al. 1986), and a decreased responsiveness in carotid and aortic tissues from rats (Fregly et al. 1977; Flaim & Hsieh, 1978; Gray, 1981). During hibernation, increased responsiveness in portal vein but not the perfused hindlimb of the hedgehog (Eliassen & Helle, 1975; Wisnes et al. 1979), and in renal but not femoral artery of the woodchuck (Miller et al. 1986) have also been reported.

The present study demonstrates increased responses of the hamster renal artery to sympathetic nerve stimulation, due to enhancement of both prejunctional and postjunctional events. Hibernation and hypothermia lead to similar patterns of decreased renal plasma flow, glomerular filtration rate and free water reabsorption rate. Reduction of glomerular filtration rate in both conditions was associated with diminished cardiac output, increased plasma oncotic pressure and increased thickness of the basement membrane (see Zatzman, 1984). Our finding of increased sympathetic neurotransmission in the renal artery of hibernating and cold-exposed hamsters offers supporting evidence for reduced blood flow to the kidneys and the resulting lower glomerular filtration rate.

It is interesting that in the renal artery, where the dominant component of transmission is noradrenergic, no significant alteration in response to the sympathetic co-transmitter ATP was established. In contrast, in the femoral arterial rings, contractile responses to ATP and the sympathetic false transmitter 5-HT were augmented during hibernation. These data demonstrate supersensitivity to ATP and 5-HT that is similar to that of denervated vessels (Araki et al. 1982; Hogestatt et al. 1988). Since there was no difference between cold-exposed and euthermic controls, this supersensitivity is due to a long term effect of hibernation.

When studying hibernation, appropriate euthermic and cold controls have to be employed in order to separate the direct effects of hibernation from the effects brought about by ambient cold. An important consideration in the design of this study was the temperature at which the arterial rings were to be studied. Because our aim was to study the long-term effects of hibernation on the vasculature, effects that should not be readily reversible, it was considered acceptable to investigate responses at 37°C. An interesting and complementary issue for further study would be to investigate vascular responses at hibernating body temperature (12°C).

Quantitative analysis of concentration-response curves for NA in renal arterial preparations revealed that maximal contractile responses were significantly greater in hibernating animals than in control animals, while no significant difference was shown between the cold controls and hibernating animals. In contrast, pD2 values did not differ among the three experimental groups, possibly indicating a receptor upregulation rather than an altered efficacy of α1-adrenoceptors following cold exposure. However, in frog heart, α-receptor efficacy is greater in the cold whereas β-receptor efficacy is greater in the warm (Chiu & Chu, 1989). Bryar et al. (1983) reported that acclimatization of rats to cold results in both a decrease in α-adrenergic and an increase in β-adrenergic responsiveness in aortic vascular smooth muscle as well as a change in the biochemical events that couple activation of adrenergic receptors to changes in vasomotor tone.

Augmented response to exogenous AVP in both cold control and hibernating animals, is reported for the first time in this study. The responses to AVP exhibited supersensitivity, similar to that of sympathectomized rats (Berecek et al. 1985). Supersensitivity is a compensatory phenomenon by which excitable cells adjust to chronic changes in the net stimulus they receive (see Fleming & Westfall, 1988). The main sources of AVP are likely to be from the circulation and from endothelial cells (Lee et al. 1988; Loesch et al. 1991), although the presence of AVP in sympathetic nerves has also been reported (Hanley et al. 1984). Hence, our results may suggest that under extremely low temperatures, AVP available from neuronal and endothelial sources, as well as circulating AVP, may become scarce, leading to tissue supersensitivity when exposed to the exogenous peptide. This hypothesis is further supported by a reduction in immunoreactivity of AVP seen in intrapineal nerve fibres of the hibernating compared with non-hibernating or arousing hedgehog (Nurnberger & Krof, 1981). There was no alteration in the response to ET-1, which points to the selective nature of changes during cold exposure and/or hibernation.

Endothelial function is influenced in various disease states, in ageing and by chronic changes in perivascular nerve input. Depressed endothelium-mediated relaxation in response to methacholine after sympathetic denervation and attenuated acetylcholine-induced endothelium-dependent vasodilatation after sensory denervation has been reported (Miller & Scott, 1990). However, in this study there was no evidence for changes in endothelial function due to hibernation, since the response to ACh was unchanged among the groups of animals.

SNP spontaneously releases NO in aqueous solution, which subsequently diffuses into smooth muscle and causes non-receptor-mediated relaxation by stimulation of cGMP (Ignarro et al. 1981). No changes in response to SNP or adenosine were seen in femoral arteries of any groups tested. Similarly, the contractile response to KCl was unaffected in both renal and femoral preparations of any of the groups. These findings demonstrate that both the contractile and dilatory apparatus of vascular smooth muscle is unaffected by long term cold exposure or hibernation. It further rules out the possibility of hypertrophy, hyperplasia and partial depolarization of vascular smooth muscle.

Neuronal innervation and responsiveness to neuro/humoral substances differs considerably amongst different vascular beds (Ralevic & Burnstock, 1995). Furthermore, heterogeneity of vascular beds in response to different diseases and animal conditions is a well documented phenomenon (Milner & Burnstock, 1995). The increase in peripheral vascular resistance that is seen on entering hibernation does not occur in all vascular beds, or to the same degree. Blood is preferentially shunted to the heart, brain, lungs and adipose tissue (Wells, 1971). The difference observed between the responsiveness of renal and femoral arteries during hibernation and/or cold exposure may be a reflection of their inherent heterogeneity and the system that they supply.

Although some effects of hibernation, including increased sympathetic neurotransmission, and increased responses to NA and AVP, are to a lesser extent mimicked by cold acclimatization, a number of vascular alterations are specific to hibernation including supersensitivity to ATP and 5-HT. It is important to recognize that hibernation and cold acclimatization have different features. Hibernation occurs after a period of adjustments resulting from physiological, biochemical, and behavioural adaptation. This hypothesis is further supported by the difference in the cheek pouch and rectal temperatures of hibernating and cold-exposed hamsters (reduced by approximately a third in hibernating hamsters), suggesting that although hibernating and cold-exposed hamsters shared similar external temperature, their body temperatures were considerably different.

It is suggested that increases in responsiveness of the renal artery to sympathetic nerve stimulation, the selective supersensitivity of vasculature to sympathetic contractile agents, and lack of any dilatory alterations may account for maintenance of vascular tone and peripheral resistance during hibernation.

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