Hemolymph Acid-base Status Research Articles

Capsaicin-induced dysregulation of acid-base status in the American cockroach

Members of TRP receptor family are involved in response to acidification. Here, we determined the effect of capsaicin, one of the TRP receptor activators, on hemolymph acid-base status in the American cockroach. Periplaneta americana adult individuals were injected with lactic acid (5% or 10%) and exposed to 100 µM capsaicin solution. Hemolymph pH was measured 15 min, 1, 4, 8 and 24 h after lactic acid and capsaicin application with a glass microelectrode. The results demonstrated that cockroaches recover from acidosis within 4 h from acid injection. Capsaicin impaired the buffering capacity of insects’ hemolymph, resulting in significant drop of hemolymph pH observed even 24 h after application. Joint action of capsaicin and acidosis reveals new insight into possible mechanism of capsaicin action on TRP receptors in insects.

Impact of ocean acidification on thermal tolerance and acid–base regulation of Mytilus edulis from the White Sea

Ocean warming and acidification are two important environmental drivers affecting marine organisms. Organisms living at high latitudes might be especially threatened in near future, as current environmental changes are larger and occur faster. Therefore, we investigated the effect of hypercapnia on thermal tolerance and physiological performance of sub-Arctic Mytilus edulis from the White Sea. Mussels were exposed (2 weeks) to 390 µatm (control) and 1120 µatm CO2 (year 2100) before respiration rate (MO2), anaerobic metabolite (succinate) level, haemolymph acid–base status and intracellular pH (pHi) were determined during acute warming (10–28 °C, 3 °C over night). In normocapnic mussels, warming induced MO2 to rise exponentially until it levelled off beyond a breakpoint temperature of 20.5 °C. Concurrently, haemolymph PCO2 rose significantly > 19 °C followed by a decrease in PO2 indicating the pejus temperature (TP, onset of thermal limitation). Succinate started to accumulate at 28 °C under normocapnia defining the critical temperature (TC). pHi was maintained during warming until it dropped at 28 °C, in line with the concomitant transition to anaerobiosis. At acclimation temperature, CO2 had only a minor impact. During warming, MO2 was stimulated by CO2 resulting in an elevated breakpoint of 25.8 °C. Nevertheless, alterations in haemolymph gases (> 16 °C) and the concomitant changes of pHi and succinate level (25 °C) occurred at lower temperature under hypercapnia versus normocapnia indicating a downward shift of both thermal limits TP and TC by CO2. Compared to temperate conspecifics, sub-Arctic mussels showed an enhanced thermal sensitivity, exacerbated further by hypercapnia, indicating their potential vulnerability to environmental changes projected for 2100.

Interactive effects of hypoxia and dissolved nutrients on the physiology and biochemistry of the freshwater crayfish Paranephrops zealandicus

A key contributing factor to the decline of the at-risk freshwater crayfish Paranephrops zealandicus is the presence of hypoxia associated with eutrophication. However, there is little information regarding the effects that dissolved nutrients have on the physiological and biochemical mechanisms that this and other freshwater crayfish use to cope with lowered dissolved oxygen. In the present study, P. zealandicus were exposed to an abrupt hypoxia (1.3kPa) in the absence or presence of either ammonia (30mgL–1 total ammonia; 0.04mgL–1 as un-ionised ammonia) or nitrite (20mgL–1) over a 6-h exposure using semi-closed box respirometry. Respiratory (oxygen consumption, haemolymph oxygen partial pressure), cardiac (heart rate) and biochemical (haemolymph acid–base status, glucose and lactate; tissue glucose and lactate) end points were measured. In hypoxia alone, haemolymph oxygen partial pressure, glucose and pH fell, whereas haemolymph lactate increased. The only significant effects of nutrients on hypoxic responses were a further decline in haemolymph oxygen partial pressure and an enhanced hypoglycaemia in the presence of nitrite. These effects may be linked to a nitrite-mediated improvement in tissue perfusion. Overall, these data indicate little additional effect of the presence of nutrients on the responses of P. zealandicus to hypoxia.

Impact of ocean acidification on thermal tolerance and acid–base regulation of Mytilus edulis (L.) from the North Sea

Anthropogenic climate change confronts marine organisms with rapid trends of concomitant warming and CO2 induced ocean acidification. The survival and distribution of species partly depend on their ability to exploit their physiological plasticity during acclimatization. Therefore, in laboratory studies the effects of simulated future ocean acidification on thermal tolerance, energy metabolism and acid–base regulation capacity of the North Sea population of the blue mussel Mytilus edulis were examined. Following one month of pre-acclimation to 10°C and control CO2 levels, mussels were exposed for two weeks to control and projected oceanic CO2 levels (390, 750 and 1120μatm) before being subjected to a stepwise warming protocol between 10°C and 31°C (+3°C each night). Oxygen consumption and heart rates, anaerobic metabolite levels and haemolymph acid–base status were determined at each temperature. CO2 exposure left oxygen consumption rate unchanged at acclimation temperature but caused a somewhat stronger increase during acute warming and thus mildly higher Q10-values than seen in controls. Interestingly, the thermally induced limitation of oxygen consumption rate set in earlier in normocapnic than in hypercapnic (1120μatm CO2) mussels (25.2°C vs. 28.8°C), likely due to an onset of metabolic depression in the control group following warming. However, the temperature induced increase in heart rate became limited above 25°C in both groups indicating an unchanged pejus temperature regardless of CO2 treatment. An upper critical temperature was reached above 28°C in both treatments indicated by the accumulation of anaerobic metabolites in the mantle tissue, paralleled by a strong increase in haemolymph PCO2 at 31°C. Ocean acidification caused a decrease in haemolymph pH. The extracellular acidosis remained largely uncompensated despite some bicarbonate accumulation. In all treatments animals developed a progressive warming-induced extracellular acidosis. A stronger pH drop at around 25°C was followed by stagnating heart rates. However, normocapnic mussels enhanced bicarbonate accumulation at the critical limit, a strategy no longer available to hypercapnic mussels. In conclusion, CO2 has small effects on the response patterns of mussels to warming, leaving thermal thresholds largely unaffected. High resilience of adult North Sea mussels to future ocean acidification indicates that sensitivity to thermal stress is more relevant in shaping the response to future climate change.

Physiological responses to emersion in the intertidal green crab, Carcinus maenas (L.)

The green shore crab, Carcinus maenas, undergoes on average 6 h periods of emersion during each low-tide cycle during the summer months. Under those conditions, the crab is cut off from its normal water environment and is exposed to potential stress from a suite of environmental and physiological changes: dehydration, compromised gas exchange and resultant internal hypoxia and hypercapnia, thermal stress, and ammonia toxicity. This study examined the comprehensive responses of the green crab in water and to a 6 h emersion period laboratory simulation of a tidal cycle followed by a 1 h re-immersion period, measuring indicators of dehydration, hemolymph osmolality, oxygen uptake, hemolymph acid–base status, heart and ventilatory rate, and hemolymph ammonia and ammonia excretion. Green crabs showed physiological responses of varying magnitude to 6 h of emersion. Individuals were found in the field exclusively under rocks and large clumps of seaweed where temperatures were approximately half those of exposed surfaces and relative humidity was about twice as high as ambient air. During emersion, crabs lost less than 5% of their wet weight, and hemolymph osmolality did not increase significantly. Oxygen uptake continued in air at about 50% of the control, aquatic values; and the gills continued to be ventilated by the scaphognathite, albeit at lower rates. Hemolymph lactate concentrations increased, indicating a shift to a greater reliance on anaerobic metabolism to support energetic needs. A slight acidosis developed in the hemolymph after 1 h of emersion, but it did not increase thereafter. Ammonia concentrations in the hemolymph were unchanged, as ammonia was volatilized by the gills and excreted into the air as NH3 gas. These results show that the green crab copes with emersion by seeking refuge in microhabitats that mitigate the changes in the physical parameters of intertidal emersion. Physiologically, desiccation is avoided, cardio-respiratory processes are maintained at reduced levels, and hemolymph acid–base balance is minimally affected. Ammonia toxicity appears to be avoided by a shift to excreting NH3 gas directly or indirectly to air.

Respiratory, cardiovascular, and hemolymph acid‐base changes in the amphibious crab, Cardisoma guanhumi, during immersion and emersion

The cardiorespiratory and hemolymph acid base status of bimodal breathing crabs, Cardisoma guanhumi, was monitored during the transition from breathing air to breathing water. Upon immersion, oxygen uptake (MO2) decreased by half. Ventilatory frequency (fsc) increased more than 5 fold, causing a decrease in hemolymph carbon dioxide partial pressure (PCO2). This was nearly fully compensated for by a gradual decrease in hemolymph bicarbonate concentration ([HCO3 ‐]) over 96 hours post‐immersion. After one to two weeks of immersion, when crabs were removed from the water, oxygen uptake initially increased, but eventually returned to the initial immersed value. Heart rate was unchanged but fsc slowed dramatically. The decreased ventilation resulted in a buildup of hemolymph PCO2, causing a respiratory acidosis that was slowly compensated for by increased hemolymph [HCO3 ‐]. C. guanhumi appears to be a truly amphibious crab with respiratory and acid‐base adaptations found in both fully aquatic and fully terrestrial species.

Acid-base and respiratory responses to hypoxia in the grasshopper Schistocerca americana.

How do quiescent insects maintain constant rates of oxygen consumption at ambient PO2 values as low as 2-5 kPa? To address this question, we examined the response of the American locust Schistocerca americana to hypoxia by measuring the effect of decreasing ambient PO2 on haemolymph acid-base status, tracheal PCO2 and CO2 emission. We also tested the effect of hypoxia on convective ventilation using a new optical technique which measured the changes in abdominal volume during ventilation. Hypoxia caused a progressive increase in haemolymph pH and a decrease in haemolymph PCO2. A Davenport analysis suggests that hypoxia is accompanied by a net transfer of base to the haemolymph, perhaps as a result of intracellular pH regulation. Hypoxia caused a progressive increase in convective ventilation which was mostly attributable to a rise in ventilatory frequency. Carbon dioxide conductance ( micromol h-1 kPa-1) across the spiracles increased more than threefold, while conductance between the haemolymph and primary trachea nearly doubled in 2 kPa O2 relative to room air. The rise in trans-spiracular conductance is completely attributable to the elevations in convective ventilation. The rise in tracheal conductance in response to hypoxia may reflect the removal of fluid from the tracheoles described by Wigglesworth. The low critical PO2 of quiescent insects can be attributed (1) to their relatively low resting metabolic rates, (2) to the possession of tracheal systems adapted for the exchange of gases at much higher rates during activity and (3) to the ability of insects to rapidly modulate tracheal conductance.

Related Changes in Hemolymph Acid-Base Status, Electrolytes, and Ecdysone in Intermolt Crayfish (Procambarus clarkii) at 23 degrees C during Extracellular Acidosis Induced by Exposure to Air, Hyperoxia, or Acid

Related Changes in Hemolymph Acid-Base Status, Electrolytes, and Ecdysone in Intermolt Crayfish (Procambarus clarkii) at 23 degrees C during Extracellular Acidosis Induced by Exposure to Air, Hyperoxia, or Acid

Haemolymph acid-base status, tracheal gas levels and the control of post-exercise ventilation rate in grasshoppers

In grasshoppers, ventilation rate increases after jumping, in association with decreases in haemolymph pH and tracheal PO2 and increases in haemolymph and tracheal PCO2. Are these changes in haemolymph acid-base status or tracheal gas composition causally responsible for the increases in post-locomotion ventilation rate? To answer this question, we manipulated haemolymph acid-base status with injections into the haemocoel and independently manipulated tracheal PO2 and PCO2 with tracheal perfusions. Using a new technique, we continuously monitored ventilation rate and ventilatory pressures on virtually unrestrained insects. Changes in haemolymph acid-base status or tracheal PCO2 did not affect post-exercise ventilation rate, clearly demonstrating that the ventilatory stimulus associated with locomotion is not dependent on negative feedback from these variables. Post-exercise ventilation rate varied with tracheal PO2, with the lowest ventilation rates observed at the lowest tracheal PO2 values, a result opposite to that expected if negative feedback from internal PO2 levels were to drive the increase in ventilation rate. Particularly after activity, there was considerable heterogeneity in unperfused animals between tracheal and haemolymph PCO2, and between tracheal PCO2 in the thorax and leg, consistent with unidirectional airflow and a considerable role for diffusion gradients in the gas exchange of grasshoppers.

Acid-base status and spiracular control during discontinuous ventilation in grasshoppers

Many insects ventilate discontinuously when quiescent, exhibiting prolonged periods during which little or no gas exchange occurs. We investigated the consequences of discontinuous ventilation (DV) on haemolymph acid­base status and tested whether spiracular opening during DV is due to changes in internal gas tensions in the western lubber grasshopper Taeniopoda eques. At 15 °C, resting T. eques exhibited interburst periods of about 40 min. During the interburst period, haemolymph PCO2 rose from 1.8 to 2.26 kPa, with minimal acidification of haemolymph. Animals in atmospheres in which PCO2 was 2 kPa or below continued to exhibit DV, while atmospheres in which PCO2 was 2.9 kPa or above caused cessation of DV. These data indicate that accumulation of internal CO2 to threshold levels between 2 and 2.9 kPa induces spiracular opening in grasshoppers. In contrast to the situation in lepidopteran pupae, variation in atmospheric PO2 had no effect on interburst duration. Relative to lepidopteran pupae, the internal PCO2 of grasshoppers during DV is threefold lower, the PCO2 required for triggering spiracular opening is also threefold lower, and the open phase spiracular conductance is at least tenfold higher, demonstrating that considerable diversity exists in these aspects of insect respiratory physiology.

Effects of extreme pH on the physiology of the Australian 'yabby' Cherax destructor: acute and chronic changes in haemolymph carbon dioxide, acid-base and ionic status

Freshwater habitats throughout the world are becoming increasingly threatened by the likelihood of acidification, but little consideration has been given to the importance of severe alkalization. Acute and chronic fluctuations in haemolymph acid­base status (PCO2, CCO2 and pH), [Na+] and [Ca2+] were monitored for up to 504 h (21 days) in the Australian freshwater crayfish Cherax destructor exposed to low- and high-pH water. The importance of carapace [Ca2+] during acid exposure was assessed. Crayfish were exposed to pH 7.1, pH 4.5 and pH 8.0 water containing calcium at 500 µmol l-1 while the effect of a lower calcium concentration (50 µmol l-1) was assessed in pH 4.5 water. Cherax in acid water containing 50 µmol l-1 Ca2+ exhibited a significant decrease in CO2 content after 2 h (mean decrease 1.13 mmol l-1, venous; 1.57 mmol l-1, arterial) and large ranges in PCO2 throughout the treatment (2.4­7.3 mmHg). The overall acid­base response was a metabolic acidosis compensated by a respiratory alkalosis. The haemolymph Na+ concentration in both control (pH 7.1, 50 µmol l-1) and acid-exposed animals in lower-Ca2+ water was up to 50 % reduced compared with that in animals in pH 7.1, 500 µmol l-1 Ca2+ water. Ion regulatory mechanisms, causing a subsequent increase in haemolymph [Na+] after 288 h, were implicated as an important component in acid­base homeostasis. Crayfish in acid, low-Ca2+ water also exhibited a 3.2 mmol l-1 increase in haemolymph [Ca2+] and showed a haemolymph alkalosis compared with animals in acid water with higher [Ca2+]. At higher water [Ca2+] in pH 4.5 water (500 µmol l-1 Ca2+), the haemolymph pH of Cherax was only 0.1 unit lower than that of animals in 50 µmol l-1 Ca2+ acid water after 96 h, and both CaCO2 and CvCO2 were unchanged compared with the initial condition. As with low-Ca2+ acid-exposure, the potential haemolymph acidosis appeared largely to be compensated by respiratory alkalosis. There was a transient 31 % reduction in haemolymph [Na+], although osmolality was unchanged (control 411±7.29 mosmol kg-1). Acid­base equilibrium recovered rapidly, probably in association with changes in ion flux and the re-establishment of normal haemolymph Na+ concentration. Alkaline-exposed Cherax destructor exhibited a mixed respiratory alkalosis and metabolic acidosis. Whereas haemolymph [Ca2+] increased by 1.8 mmol l-1 after only 1 h, haemolymph Na+ levels increased by 36 % after 2 h, possibly as part of a net H+ loss from the haemolymph. Increased HCO3-/Cl- exchange could contribute to the 4.3 mmol l-1 decrease in haemolymph CO2 level after 0.5 h of alkaline exposure. The responses of Cherax to extreme pH are different from those of the European and North American crayfish species studied to date.

Acid-Base Variables in Malpighian Tubule Secretion and Response to Acidosis

1. Malpighian tubule fluid from Schistocerca gregaria adults, starved for 1 day, was collected in situ by cannulation of the gut, both before and after injecting 10 μmol of HCl or NaCl into the haemocoel. 2. Haemolymph pH at the neck remained depressed by 0.3 units for at least 6 h in HCl- as compared to NaCl-injected locusts. A lower haemolymph pH persisted near the acid injection site for several hours. 3. The pH of tubule fluid remained about 0.5 units more acid than haemolymph under all conditions. Thus, net tubular acid secretion was proportional to haemolymph acid-base status. 4. The greater acidity of tubular fluid after acid injection was associated with lower estimated bicarbonate concentrations and higher Pcoco2 without any change in total CO2 when compared to controls. 5. The combined contribution of bicarbonate, phosphate and urate to total buffering capacity of tubular fluid was estimated to be 75°, with bicarbonate responsible for 55° of the total. 6. The maximum rate of acid removal by all Malpighian tubules of starved locusts, including H+ trapped in ammonium ions, was calculated to be very small in relation to the acid load injected into the haemocoel Note: To whom reprint requests should be directed.

Metabolic response of the grass shrimp Palaemonetes kadiakensis Rathbun, to acute exposure of sublethal changes in pH

The effects of sublethal changes in environmental pH on the grass shrimp Palaemonetes kadiakensis were determined using changes in metabolic rate and critical oxygen concentration as indicators of stress. Oxygen consumption was measured using a computer automated intermittent flow respirometer which allowed multiple sampling of a single individual without exposing it to prolonged oxygen stress. Metabolic rate and critical oxygen concentration were determined using a quadratic model with plateau nonlinear regression procedure. Sublethal changes in pH (6.5, 9.0) did not have a significant effect on metabolic rate. Temporary disturbances in hemolymph acid-base status caused by acute pH stress, may not elicit a metabolic response when not accompanied by necrosis of the gill epithelium. There was an effect of pH on critical oxygen concentration, with pH 6.5 significantly higher than the control (pH 7.8), and pH 9.0 significantly lower than the control. Shifts in critical oxygen concentration were attributed to changes in blood oxygen affinity caused by hemolymph acidosis or alkalosis.

Ventilatory frequency and haemolymph acid-base status during short-term hypercapnia in the locust, Schistocerca nitens

In this study, I investigated the role of the locust ventilatory system in haemolymph acid-base regulation during short-term (50 min) exposure to elevated environmental carbon dioxide. In American locusts, Schistocerca nitens, abdominal pumping frequency increased linearly with inspired P CO 2 ( P 1CO 2) to a P 1CO 2 of 107 mmHg (1 mmHg = 133.3 Pascals). Ventilatory frequency was strongly correlated with haemolymph acid-base variables, consistent with the hypothesis that changes in ventilatory frequency were caused by changes in haemolymph pH or P CO 2 . As inspired P CO 2 increased, the partial pressure gradient for carbon dioxide from haemolymph to air decreased, reducing the changes in haemolymph pH. Slopes of in vivo and in vitro buffer lines were statistically indistinguishable, suggesting that changes in haemolymph pH in vivo were due primarily to changes in haemolymph P CO 2 . The results of this study indicate that the locust ventilatory system plays the dominant role in limitation of changes in haemolymph pH during short-term hypercapnia.

Temperature Effects on Haemolymph Acid-Base Status in vivo and in vitro in the Two-Striped Grasshopper Melanoplus Bivittatus

ABSTRACT In this study, I examine the effect of temperature on haemolymph acid-base status in vivo and in vitro in the two-striped grasshopper Melanoplus bivittatus. Melanoplus bivittatus experience wide (up to 40 °C) diurnal body temperature fluctuations in the field, but maintain body temperature relatively constant during sunny days by behavioural thermoregulation. Haemolymph pH was statistically constant (7·12) between 10 and 25°C, but decreased by –0·017 units °C− 1 from 25 to 40 °C. Relative alkalinity and fractional protein dissociation were conserved only at body temperatures at which feeding and locomotory activity occur, above 20°C. Haemolymph total CO2 (Ctot) increased from 10 to 20°C and decreased from 20 to 40°C. Haemolymph increased from 10 to 20°C and was statistically constant between 20 and 40°C. Carbonic acid pKapp in haemolymph was 6·122 at 35 °C, and decreased with temperature by –0·0081 units °C− 1. Haemolymph buffer value averaged – 35mequivl− 1pHunit− 1. Haemolymph pH changes with temperature were small (less than –0·004 units °C− 1) in vitro at constant . Therefore, passive physicochemical effects cannot account for the pattern of acid-base regulation in vivo. The temperature shift from 10 to 20°C was accompanied by a net addition of 4·2–0·2 mmol I− 1 of bicarbonate equivalents to the haemolymph. The temperature shift from 20 to 40°C was accompanied by a net removal of 10–14 mmol I− 1 of bicarbonate equivalents from the haemolymph. Haemolymph acid-base regulation in vivo during temperature changes is dominated by active variation of bicarbonate equivalents rather than by changes in as observed for most other air-breathers.

Acid-base balance and blood gas changes in the fresh water field crab, Barytelphusa guerini, on exposure to organic and inorganic lead.

The acid-base status of crustacean haemolymph depends on various environmental and physiological factors. Acid base status of the haemolymph is known to be influenced by temperature, salinity, strenuous activity and moulting. The studies on the acid-base regulation of the fresh water crabs are meager. The acid-base changes in fishes during environmental stress conditions like acid stress and zinc toxicity had been reported. But the effect of environmental pollutants like the heavy metals on the acid-base regulation of the fresh water crabs have not been previously reported. The haemolymph of the fresh water crab was found to accumulate high amounts of lead on exposure to organic and inorganic lead. Hence the present investigation has been undertaken to study the haemolymph acid-base status on exposure to subtoxic levels of organic and inorganic lead.

Gas Exchange, Hemolymph Acid-Base Status, and the Role of Branchial Water Stores during Air Exposure in Three Littoral Crab Species

O₂ uptake and hemolymph acid-base status, together with branchial water volume, CO₂ content, and titratable alkalinity, were measured in three species of intertidal crabs. In Pachygrapsus crassipes, a grapsid crab that actively moves between air and water, O₂ uptake increased on emersion. In Eurytium albidigitum, a mud-burrowing xanthid crab that is air exposed by tidal action, O₂ uptake declines dramatically on emersion. There is no significant lactate production by either crab following emersion. An emersion-induced respiratory acidosis was fully compensated in P. crassipes and another grapsid, Hemigrapsus nudus, but uncompensated in E. albidigitum. Branchial water volume 10 min after emersion was 0.013 ml/g crab weight in P. crassipes and 0.072 ml/g in E. albidigitum. The CO₂ content of branchial water in P. crassipes increased rapidly during air exposure and was accompanied by an increase in titratable alkalinity (TA). The CO₂ content of branchial water in E. albidigitum remained constant for at least...

CHANGES IN THE HEMOLYMPH ACID-BASE STATE OF THE SHORE CRAB,CARCINUS MAENAS, EXPOSED TO SIMULATED TIDEPOOL CONDITIONS

High-tide and low-tide conditions were established in an artificial pool containing algae by alternately circulating seawater and leaving it stagnant for successive 6-h periods. Diurnal cyclical changes of water temperature and partial pressures of oxygen and carbon dioxide were almost the same as in natural tidepools. Hemolymph acidbase status (pH, carbon dioxide partial pressure, and bicarbonate concentration) was determined during day and night low-tide periods in crabs (Carcinus maenas) acclimated to artificial pool conditions. Except at water Po2 below 20 Torr where the animals breathed air and developed a partially compensated respiratory acidosis, acidbase disturbances induced by changes of respiratory gases were much less than those observed in single-factor laboratory experiments, mainly because of counteracting influences of oxygen and carbon dioxide. This resulted in a decrease of hemolymph pH at increasing water temperature of –0.016 pH unit °C-1, consistent with the imidazole alphastat hypothesis.

The role of branchial ventilation in hemolymph acid-base changes in the shore crabCarcinus maenas during hypoxia

1. When exposed to hypoxia in the ambient medium, the crabCarcinus maenas increased and sustained high levels of branchial ventilation for up to 70 h (Fig. 1). 2. The role of hyperventilation in establishing changes in hemolymph acid-base status during hypoxia was investigated. Hemolymph pH and\(P_{CO_2 } \) changes in hypoxic crabs (Figs. 2 and 4) were compared with crabs whose branchial chambers were artificially hyperventilated in normoxia by siphoning water through a mask attached to the carapace (Fig. 3). 3. Hyperventilation alone does not account for the observed alkalosis during hypoxia. It is suggested that changes in both CO2 production and ventilation may be responsible for altering hemolymph acid-base status. 4. Artificial hyperventilation in normoxic crabs resulted in a respiratory alkalosis which is fully compensated after 16 hours by a metabolic acidosis (Fig. 3). 5. Reduction of the hemolymph bicarbonate pool during hypoxia did not interfere with the ability of crabs acclimated to low salinity to regulate hemolymph chloride ion concentration (Fig. 5).